1. Manual coring
<ol>
	<li>Assemble the increment corer and select the coring position on the stem of a tree depending on the research question (e.g., for geomorphic reconstructions, parallel to the direction of mechanical stress; for age determination, as low down as possible). 
	NOTE: Always take two cores from each stem, preferably in the opposite direction.</li>
	<li>After selecting the coring position, place the corer at a right angle relative to the direction of growth of the stem.</li>
	<li>Place a pusher at the rear end of the corer to stabilize it while drilling.</li>
	<li>Achieve a stable position and lean against the pusher to apply pressure to the cutting edge.</li>
	<li>Turn the handle of the corer with both hands until the threaded part of the drill is fully turned into the stem.</li>
	<li>Release the pressure and remove the pusher.</li>
	<li>Start turning the handle of the corer with both hands until the corer has reached or drilled through the pith. Check this by holding the extractor (which has the same length as the corer) on the handle aside the stem.</li>
	<li>Take the extractor with the open side on top and insert it fully into the corer. Turn the corer backward (one full turn) to break the core from the stem. Pull the extractor out of the corer.</li>
	<li>Remove the core from the extractor and store it in a paper straw.</li>
	<li>Remove the corer from the stem and store it in the handle.</li>
</ol>
2. Coring with a cordless drill
<ol>
	<li>Take the cordless drill equipped with a torque booster and add the special adapter for the increment corer developed at WSL.</li>
	<li>Place the increment corer into the adapter on the torque booster and select the position for coring on the stem of a tree depending on the research task (see step 1.1).</li>
	<li>After selecting the coring position, place the corer at a right angle relative to the direction of growth of the stem.</li>
	<li>Achieve a stable position, hold the cordless drill tight, and apply pressure on the cutting edge.</li>
	<li>Start the cordless drill, turning slowly until the threaded part of the drill is fully cored into the stem, then increase the speed until the corer reaches or drills through the pith. 
	NOTE: The depth can be checked as explained in step 1.7.</li>
	<li>Remove the cordless drill from the corer, place the handle on it, and use the extractor to remove the core as explained in step 1.8.</li>
	<li>Store the increment core in a paper straw.</li>
	<li>Remove the handle, place the cordless drill on the corer and remove the corer from the stem.</li>
</ol>
3. Sharpening the cutting edge of increment corers
<ol>
	<li>Using the WSL sharpening support
	<ol>
		<li>Take the newly designed holder and place it on the ground.</li>
		<li>Place the cordless drill, including the increment corer, at the designated support points and close the mounting bracket to fix the cordless drill.</li>
		<li>Start the cordless drill by fixing the Teflon block on the starter button and let it run. Take the conical grinding stone and grind the inside of the cutting edge with it. 
		NOTE: The contact angle depends on the inside of the cutting edge. The grinding stone should have full contact with the inner sidewall, reaching from the cutting edge until the inner widening of the corer.</li>
		<li>Take the rectangular grindstone and grind the outside of the cutting edge to deburr it. 
		NOTE: This is necessary to remove the previously formed burr on the edge by grinding it from the inside and to finally sharpen the edge.</li>
		<li>Remove the Teflon block from the starter button to stop the drill, open the mounting bracket to release the cordless drill, and take the device out of the holder.</li>
	</ol>
	</li>
	<li>Checking the sharpness of the cutting edge
	<ol>
		<li>Remove the increment corer from the adapter of the cordless drill.</li>
		<li>Place a sheet of paper on the wooden board of the sharpening support.</li>
		<li>Place the cutting edge of the corer on the paper while holding the corer vertically.</li>
		<li>Turn the corer while holding it vertically without applying pressure to the corer-only the weight of the corer should be pressing on the paper.</li>
		<li>Lift the corer and check whether a round piece of the paper has remained inside the cutting edge of the corer. If so, the corer is sharp. If not, repeat the sharpening procedure (step 3.1). Repeat the entire procedure (steps 3.1 and 3.2) if the outside of a core is not smooth.</li>
	</ol>
	</li>
</ol>
5. Cutting micro sections of entire increment cores using a water-soluble tape
<ol>
	<li>Place a long glass slide next to the microtome and add some water in the middle of the slide along its entire length.</li>
	<li>Place the core in the sample holder of a core microtome. 
	NOTE: To cut the section as a true cross section, make sure that the direction of the fibers is upright.</li>
	<li>Lift the sample holder until the core almost touches the edge of the blade. Pull the blade over the core to cut off the top.</li>
	<li>Place the knife at the beginning of the core again, lift the sample about 10 &#181;m, and repeat the cutting procedure until a plane surface of at least 2 mm width is obtained.</li>
	<li>Add a corn starch solution on the cut surface using a brush21.</li>
	<li>Use a piece of cloth to remove the surplus solution from the top of the core.</li>
	<li>Cut a strip of water-soluble tape to the same length as the core; place one side of the tape to the start of the core, with an overlap of approximately 1 cm, with the beginning of the core facing the blade of the microtome.</li>
	<li>Attach the tape to the surface of the core by stroking the tape on the surface using a finger.</li>
	<li>Lift the sample in the microtome by 15-20 &#181;m, lift the overlapping piece of the tape a little, and place the microtome blade at the edge of the core.</li>
	<li>Cut the section while holding the end of the tape.</li>
	<li>Take the tape with the thin section stuck on it and place it with the cut facing down on the line of water of the glass slide prepared in step 5.1.</li>
	<li>After approximately 10 s, start removing the tape using tweezers, by holding the tape on one side and lifting it up, while taking care that the section remains on the glass slide.</li>
	<li>To produce a permanent slide of this section, follow the standard procedures22.</li>
</ol>

<ol>
	<li>B&#252;ntgen, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scrutinizing+tree-ring+parameters+for+Holocene+climate+reconstructions.">Scrutinizing tree-ring parameters for Holocene climate reconstructions.</a> Wiley Interdisciplinary Reviews: Climate Change. , 778 (2022).</li><li>Hadad, M. A., Gonz&#225;lez-Reyes, &. #. 1. 9. 3. ;., Roig, F. A., Matskovsky, V., Cherubini, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tree-ring-based+hydroclimatic+reconstruction+for+the+northwest+Argentine+Patagonia+since+1055+CE+and+its+teleconnection+to+large-scale+atmospheric+circulation.">Tree-ring-based hydroclimatic reconstruction for the northwest Argentine Patagonia since 1055 CE and its teleconnection to large-scale atmospheric circulation.</a> Global and Planetary Change. 202, 103496 (2021).</li><li>Shen, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+climate+on+the+tree+ring+density+and+weight+of+Betula+ermanii+in+a+cool+temperate+forest+in+central+Japan.">Effects of climate on the tree ring density and weight of Betula ermanii in a cool temperate forest in central Japan.</a> Trees. , 1-9 (2022).</li><li>Wang, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acoustic+measurements+on+trees+and+logs:+a+review+and+analysis.">Acoustic measurements on trees and logs: a review and analysis.</a> Wood Science and Technology. 47 (5), 965-975 (2013).</li><li>Downes, G. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Application+of+the+IML+Resistograph+to+the+infield+assessment+of+basic+density+in+plantation+eucalypts.">Application of the IML Resistograph to the infield assessment of basic density in plantation eucalypts.</a> Australian Forestry. 81 (3), 177-185 (2018).</li><li>Tomczak, K., Tomczak, A., Jelonek, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Measuring+radial+variation+in+basic+density+of+pendulate+oak:+comparing+increment+core+samples+with+the+Iml+power+drill.">Measuring radial variation in basic density of pendulate oak: comparing increment core samples with the Iml power drill.</a> Forests. 13 (4), 589 (2022).</li><li>Piene, H., D'Amours, J., Bray, A. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spruce+budworm+defoliation+and+growth+loss+in+young+balsam+fir:+estimation+of+volume+growth+based+on+stem+analysis+and+increment+cores+at+breast+height.">Spruce budworm defoliation and growth loss in young balsam fir: estimation of volume growth based on stem analysis and increment cores at breast height.</a> Northern Journal of Applied Forestry. 13 (2), 73-78 (1996).</li><li>Phipps, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Collecting,+Preparing,+Crossdating,and+Measuring+Tree+Increment+Cores.">Collecting, Preparing, Crossdating,and Measuring Tree Increment Cores.</a> US Department of the Interior, Geological Survey. , (1985).</li><li>Schweingruber, F. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Tree Rings and Environment: Dendroecology. , (1996).</li><li>Toole, E. R., Gammage, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Damage+from+increment+borings+in+bottomland+hardwoods.">Damage from increment borings in bottomland hardwoods.</a> Journal of Forestry. 57 (12), 909-911 (1959).</li><li>Grissino-Mayer, H. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+manual+and+tutorial+for+the+proper+use+of+an+increment+borer.">A manual and tutorial for the proper use of an increment borer.</a> Tree-Ring Research. 59 (2), 63-79 (2003).</li><li>Wunder, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Does+increment+coring+enhance+tree+decay?+New+insights+from+tomography+assessments.">Does increment coring enhance tree decay? New insights from tomography assessments.</a> Canadian Journal of Forest Research. 43 (8), 711-718 (2013).</li><li>Helcoski, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=No+significant+increase+in+tree+mortality+following+coring+in+a+temperate+hardwood+forest.">No significant increase in tree mortality following coring in a temperate hardwood forest.</a> Tree-Ring Research. 75 (1), 67-72 (2019).</li><li>Hall, A. A., Bloomberg, W. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+power-driven+increment+borer.">A power-driven increment borer.</a> The Forestry Chronicle. 60 (6), 356-357 (1984).</li><li>Scott, J. H., Arno, S. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Using+a+power+increment+borer+to+determine+the+age+structure+of+old-growth+conifer+stands.">Using a power increment borer to determine the age structure of old-growth conifer stands.</a> Western Journal of Applied Forestry. 7 (4), 100-102 (1992).</li><li>Krottenthaler, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+power-driven+increment+borer+for+sampling+high-density+tropical+wood.">A power-driven increment borer for sampling high-density tropical wood.</a> Dendrochronologia. 36, 40-44 (2015).</li><li>Caetano-Andrade, V. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Advances+in+increment+coring+system+for+large+tropical+trees+with+high+wood+densities.">Advances in increment coring system for large tropical trees with high wood densities.</a> Dendrochronologia. 68, 125860 (2021).</li><li>Edwards, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intra-annual+climate+anomalies+in+northwestern+North+America+following+the+1783-1784+CE+Laki+eruption.">Intra-annual climate anomalies in northwestern North America following the 1783-1784 CE Laki eruption.</a> Journal of Geophysical Research: Atmospheres. 126 (3), 033544 (2021).</li><li>Zhirnova, D. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+495-year+wood+anatomical+record+of+Siberian+stone+pine+(Pinus+sibirica+Du+Tour)+as+climatic+proxy+on+the+timberline.">A 495-year wood anatomical record of Siberian stone pine (Pinus sibirica Du Tour) as climatic proxy on the timberline.</a> Forests. 13 (2), 247 (2022).</li><li>Maeglin, R. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Increment Cores: How to Collect, Handle, and Use Them. 25, (1979).</li><li>G&#228;rtner, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+technical+perspective+in+modern+tree-ring+research+-+how+to+overcome+dendroecological+and+wood+anatomical+challenges.">A technical perspective in modern tree-ring research - how to overcome dendroecological and wood anatomical challenges.</a> Journal of Visualized Experiments. (95), e52337 (2015).</li><li>G&#228;rtner, H., Banzer, L., Schneider, L., Schweingruber, F. H., Bast, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preparing+micro+sections+of+entire+(dry)+conifer+increment+cores+for+wood+anatomical+time-series+analyses.">Preparing micro sections of entire (dry) conifer increment cores for wood anatomical time-series analyses.</a> Dendrochronologia. 34, 19-23 (2015).</li><li>Rodriguez, D. R. O., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exploring+wood+anatomy,+density+and+chemistry+profiles+to+understand+the+tree-ring+formation+in+Amazonian+tree+species.">Exploring wood anatomy, density and chemistry profiles to understand the tree-ring formation in Amazonian tree species.</a> Dendrochronologia. 71, 125915 (2022).</li><li>G&#228;rtner, H., Farahat, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cambial+activity+of+Moringaperegrina+(Forssk.)+Fiori+in+arid+environments.">Cambial activity of Moringaperegrina (Forssk.) Fiori in arid environments.</a> Frontiers in Plant Science. 12, 760002 (2021).</li><li>von Arx, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Q-NET-a+new+scholarly+network+on+quantitative+wood+anatomy.">Q-NET-a new scholarly network on quantitative wood anatomy.</a> Dendrochronologia. 70, 125890 (2021).</li><li>Seftigen, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prospects+for+dendroanatomy+in+paleoclimatology-a+case+study+on+Picea+engelmannii+from+the+Canadian+Rockies.">Prospects for dendroanatomy in paleoclimatology-a case study on Picea engelmannii from the Canadian Rockies.</a> Climate of the Past. 18 (5), 1151-1168 (2022).</li><li>Matulewski, P., Buchwal, A., G&#228;rtner, H., Jagodzi&#324;ski, A. M., &#268;ufar, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Altered+growth+with+blue+rings:+comparison+of+radial+growth+and+wood+anatomy+between+trampled+and+non-trampled+Scots+pine+roots.">Altered growth with blue rings: comparison of radial growth and wood anatomy between trampled and non-trampled Scots pine roots.</a> Dendrochronologia. 72, 125922 (2022).</li><li>Prislan, P., del Castillo, E. M., Skoberne, G., &#352;penko, N., Gri&#269;ar, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sample+preparation+protocol+for+wood+and+phloem+formation+analyses.">Sample preparation protocol for wood and phloem formation analyses.</a> Dendrochronologia. 73, 125959 (2022).</li><li>Heinrichs, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pocket-sized+sharpender+for+increment+borers.">Pocket-sized sharpender for increment borers.</a> Journal of Forestry. 62, 653 (1964).</li><li>Schneider, L., G&#228;rtner, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Additive+manufacturing+for+lab+applications+in+environmental+sciences:+pushing+the+boundaries+of+rapid+prototyping.">Additive manufacturing for lab applications in environmental sciences: pushing the boundaries of rapid prototyping.</a> Dendrochronologia. 76, 126015 (2022).</li></ol>

The authors declare that they have no competing financial interests.

The significant inclusion of wood anatomy in dendroecological studies23,24,&#160;as well as an intensified exchange among scientists specialized in tree-ring research and wood anatomists25, has opened a wide field of new and in-depth analyses of past environmental conditions. These new studies have opened up new possibilities and questions, but have also given rise to new problems.
The fast development of this new era of &#34;dendroanatomy&#34; requires a high number of samples, which is definitely supported by the use of a cordless drill as explained before. In addition to the fact that taking cores with the drill is not at all exhausting, it saves a lot of time. Although the results presented in this paper imply sampling possibilities that are six times faster than manual coring, it is a test for single cores. Nevertheless, during regular sampling (one person coring, with one coding and storing the cores), we managed to core 24 spruce trees (two cores of full length each), with stem diameters of approximately 80 cm, within 1.5 h. This is an average of &#60;2 min for one core, including storage, packing, and moving to the next tree.
The fast handling of the entire process is supported by the fact that the newly designed adapter for increment corers can be used without the need to fix the corer inside the adapter with a screw or comparable closures. As a result, changing the drill to the handle of the corer to break and extract the core is fast and easy. The adapter is designed so that one can even pull the corer out while drilling back in case the stem is rotten, or (as is common with some increment corers) if the thread does not grab when turning back and the corer does not move out.
However, it should be noted that when removing the corer from the stem, one needs to tilt the adapter slightly so that it can be successfully pulled without the drill slipping off (protocol step 2.8). The increasing demand for tree-ring studies to create long chronologies based on anatomical proxies19,26 has required the preparation of micro sections from increment cores, cut in pieces before preparation or cut as entire micro sections22. Although the quality of up to 40 cm long micro sections is still not always comparable to short sections (e.g., the variable angle of the cells in their vertical extension frequently hampers cell wall measurements), they can be used to identify and date specific growth reactions as the occurrence of reaction wood or blue rings27 (Figure 6).
Consequently, the quality of the samples is a basic prerequisite for successful preparation and further analyses of anatomical structures. This demand requires more caution regarding the sharpness of the sampling campaign when taking increment cores. As a consequence, preparing micro sections can be very time-consuming and labor-intensive, and sometimes even impossible, if the specimens are not embedded beforehand28.
Sharpening the cutting edge of an increment corer by hand requires a lot of practice and experience, in order to grind the edge evenly all around by hand without any support. The ability of using the new drill mount for sharpening increment cores enables even users inexperienced in sharpening to sharpen the cutting edge of their corers in the field. The fact that this now can be done quickly will increase the quality of samples taken in future.
Although the use of the new equipment shows clear advantages for subsequent processing of the cores, the cordless drill could also be combined with small devices for sharpening, developed and presented almost 40 years ago20. Maeglin20 presented construction details of a modification of &#34;Goodchild&#39;s borer sharpener&#34; made of wood and metal29. Nowadays, this device can be modeled and printed in a 3D printer without any problems30. One would only need to create a detailed 3D model of the sharpener to print the single parts and assemble it to use in the field. The possibilities for improvement have not yet been exhausted and we are sure that this publication will inspire many colleagues to further develop the tools presented here. A yet unsolved obstacle is the fact that one needs to remove the drill and add the handle of the corer to extract the core.
The final step of cutting micro sections of entire increment cores22 is still a tricky issue. The application of the water-soluble tape, as described earlier, supports the process by stabilizing the section when cutting and placing it on the glass slide. Nevertheless, this procedure still requires of the user to have a high level of experience.

Dendroecological research is based on various characteristics of growth rings in trees, both annual and otherwise. &#34;Precursory&#34; discipline dendrochronology was established using ring-width variations as a parameter to simply date the rings and, as a result, establish long chronologies. Therefore, manifold other characteristics, such as density variations, isotopic concentrations, or wood anatomical characteristics, are used to correlate single rings or their structure and content to environmental parameters to better understand the impact of environmental conditions on tree growth over time.
Dendroecology, as well as dendroclimatology, has gained in importance in environmental research, mainly in reconstructing past climate conditions1,2,3. For this, the rings of countless trees have to be analyzed in detail. Although some techniques exist to determine tree-ring width and density (e.g., by acoustic wave technology4 or drilling resistance5,6), to date, there is no reliable &#34;non-destructive&#34; method to extract the characteristics of rings from trees. For very detailed analyses of ring characteristics within a tree, or to estimate basal area increment, it would be best to cut disks from the trees of interest7. This would require cutting down all potential trees of interest for specific analyses. Bearing in mind the huge number of trees analyzed worldwide each year, this sampling strategy is not practicable. Regardless of wasting an incredible amount of resources, this strategy is simply too expensive. Due to this, the use of increment corers has been established as a standard sampling technique in tree-ring research8. The use of increment corers allows for a minimally invasive extraction of wood cores from stems, starting from the bark and reaching (in optimal cases) the pith of the tree9.
Although coring causes an injury to the stem-a hole with a diameter of ~1 cm-trees are able to close this wound through increased wood formation in the vicinity of the core hole. A disadvantage, apart from the hole itself, is the occurrence of a &#34;compartmentalization zone&#34;, an area around the hole where the cells are filled by phenols to prevent the potential spread of fungi starting from the hole10,11. To our knowledge, there is still no evidence that increment coring causes a significant increase of tree decay frequency, at least in undisturbed high-elevation forest stands for Picea abies12 and several hardwood species in a temperate forest13.
Although this sampling standard has been applied for decades all over the world, some problems still remain. One of these is the fact that the cores have to be taken by hand without any mechanical support, which takes a lot of time and is quite exhausting after a while. To ease sampling, several (more or less practicable) strategies have been tested, such as the use of chainsaws equipped with a corer instead of the chain14,15,16,17. The use of chainsaws was preferred to drills because the latter were not powerful enough; however, this idea did not catch on due to the large weight of the chainsaw and the fuel required.
In recent years, wood anatomical techniques have evolved significantly and been integrated into dendroecological studies18,19. However, the ability to analyze wood anatomical parameters over long periods by cutting micro sections from increment cores resulted in unexpected problems. Frequently, the micro sections taken from cores broke into little pieces, which made it impossible to produce coherent cuts (Figure 1). This problem was caused by the manual technique of coring trees and unsharp corers. The mechanical stress exerted on the wood while coring resulted in micro cracks within the core. These micro cracks were never noticed during macroscopic examination of the increment cores, and therefore never presented a problem.
Manual coring is done by placing the handle on the rear end of the corer, pressing the tip with the thread to the stem, and starting to turn the handle until the corer has pierced a little over half the diameter of the stem. While doing this, the tip of the corer is (obviously) fixed in the stem, but the rear end of the corer turned by the handle is always moving sideward or up and down, at least until the drill head is fully screwed into the trunk, giving more guidance and stability to the corer. As a result of the high pressure and the movement of the corer, the increment cores are distorted frequently in the outermost ~5 cm (Figure 1). Even if the friction while turning is reduced to a minimum, another process is exerting stress on the increment core inside the corer. Manual coring does not allow a continuous movement of the cutting edge of the corer inside the stem. One can do a maximum of one full turn, before having to stop to change the grip, and then continue drilling. Each time the rotation restarts, the core is slightly twisted until the friction is overcome and the drill rotates again. These mechanical stresses potentially cause microscopic cracks in the structure of the cores.
This mechanical stress is even increased when the cutting edge of the corer is not sharp. A visible sign for an unsharp corer is an uneven core surface, showing lots of cracks along its entire extension20 (Figure 2). The frequency of sharpening depends on the density of the trees to be cored and the minerals or sand present in the bark of the tree to be cored. On a general note, one should not assume that new corers are sharp. To date, sharpening a corer is almost never done in the field due to the difficulty of it, since this has to be done by hand and needs a lot of experience11,20.
To summarize, manual coring and unsharp cutting edges both result in micro cracks occurring in cores taken. To date, these problems have not been analyzed systematically, nor have attempts been made to find solutions. This paper presents a protocol to overcome these obstacles by comparing the manual coring technique to the application of a new technique. We propose using a cordless drill equipped with a special adapter for an increment corer. We present to which extent problems are minimized when coring a tree, as well as the effect of continuous, mechanical coring on the preparation of long micro sections. This protocol includes the preparation of long micro sections using a water-soluble tape as a supporting aid and a procedure to sharpen corers in the field.

<table><tbody><tr><td>BS 18 LTX-3 BL QI</td><td>Metabo</td><td></td><td>Cordless drill</td></tr><tr><td>Core-microtome</td><td>WSL</td><td></td><td>Microtome to cut micro sections from increment cores</td></tr><tr><td>Drill adapter for increment corer</td><td>WSL</td><td></td><td>Adapter to fix the increment corer on the cordless drill</td></tr><tr><td>Increment corer</td><td>Hagl&amp;ouml;ff</td><td></td><td>40cm increment corer</td></tr><tr><td>Power X3</td><td>Metabo</td><td></td><td>Torque amplifyer</td></tr><tr><td>Sharpening support board</td><td>WSL</td><td></td><td>Board to attach the cordless dril to sharpen the cutting edge ofd the corer</td></tr><tr><td>Water-soluble tape 5414, transparent 3/4IN</td><td>3M</td><td></td><td>Transparent tape to support cutting long sections</td></tr></tbody></table>

advanced workflow for taking high-quality increment cores - new techniques and devices

In dendroecological research, precise dating of each single growth ring is a basic requirement for all studies, focusing on ring-width variations only, chemical or isotope analyses, or wood anatomical studies. Independent of the sampling strategy for a certain study (e.g., climatology, geomorphology), the way samples are taken is crucial for their successful preparation and analyses.
Until recently, it was sufficient to use a (more or less) sharp increment corer to obtain core samples that could be sanded for further analyses. Since wood anatomical characteristics can be applied to long time series, the need to obtain high-quality increment cores has taken on a new meaning. Essentially, the corer needs to be sharp(ened) when used. When coring a tree by hand, there are some problems in handling the corer, resulting in the hidden occurrence of micro cracks along the entire core: When starting to drill by hand, the drill bit is strongly pressed against the bark and the outermost ring until the thread has fully entered the trunk. At the same time, the drill bit is moved up and down as well as sideward. Then, the corer is drilled all the way into the trunk; however, it is necessary to stop after each turn, change the grip, and turn again. All these movements, as well as the start/stop-coring, puts mechanical stress on the core. The resulting micro cracks make it impossible to create continuous micro sections, as they fall apart along all these cracks.
We present a protocol to overcome these obstacles by applying a new technique using a cordless drill to minimize these problems when coring a tree, as well as its effect on the preparation of long micro sections. This protocol includes the preparation of long micro sections, as well as a procedure to sharpen corers in the field.

When comparing the manual coring procedure to the use of a cordless drill, the advantages of the latter are obvious. We compared coring spruce trees (Picea abies (L.) H. Karst.) with a stem diameter at a breast height of 60-80 cm. We used 5 mm corers, with a length of 40 cm, for all cores taken, and drilled the full length of the corer into the stem. When taking cores manually, the full procedure of taking a core and removing the corer from the tree again took an average of ~6 min. When repeating this using the cordless drill equipped with a torque booster, the entire procedure took an average of only 1 min. In addition to the fact that coring with the cordless drill is not at all exhausting, none of the cores were deformed, due to pressure exerted on the cutting edge during the first phase of drilling until the thread is completely inside the stem. As soon as the thread is inside the stem, the corer is more or less stabilized, and potential up- and downward movements are minimized (Figure 3).
As soon as the first core was no longer smooth on the outside but showed scratches and cracks, as in Figure 2, sharpening of the cutting edge was required. Because the cordless drill can be fixed, as it is used for coring (i.e., including the adapter and the increment corer; Figure&#160;4), the sharpening procedure is also quite fast. With a little practice, sharpening takes no more than 5 min. As soon as the cut paper sticks inside the corer, sampling can be continued. The resulting cores are smooth without any scratches or cracks. The cores taken with the aid of the cordless drill have a lower probability of showing micro cracks; this is a precondition to cut micro sections of entire increment cores. The application of the water-soluble tape (Figure 5) made the handling of the long and fragile sections easier, since the tape protects the thin section from tearing while taking it off the blade and placing it on the glass slide. This procedure saves time in the lab and improves the quality of the micro sections, because the tape glue stabilizes the cell walls while cutting in addition to the non-Newtonian fluid (corn starch solution; see protocol step 5.5).
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/64747/64747fig01.jpg" /> 
Figure 1: Increment corer. (A) Increment corer, as used for manual coring, and a magnified view of the thread and the cutting edge. (B) Distorted increment core due to high pressure exerted to the wood at the beginning of manual coring. (C) Micro section of a part of an increment core fragmented due to micro cracks. Scale bar = 0.5 cm. (D-F) Photos indicating the procedure of coring when using a cordless drill. No high pressure is needed to start coring (D,E), the handle can easily be used to extract the core (F), and the drill is extracted directly thereafter (G). <a href="https://www.jove.com/files/ftp_upload/64747/64747fig01large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/64747/64747fig02.jpg" /> 
Figure 2: Increment core showing scratches and cracks on the outside due to the use of an unsharp corer. Scale bar = 0.5 cm. <a href="https://www.jove.com/files/ftp_upload/64747/64747fig02large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/64747/64747fig03.jpg" /> 
Figure 3: Increment cores and related micro sections. (A) Straight core taken with a sharp corer and a cordless drill. (B) Distorted core taken manually with an unsharp corer. (C) Continuous section of a Pinus sylvestris core sampled with a sharp corer. (D) Section of a Larix decidua core broken in pieces due to using an unsharp corer. Scale bars = 0.5 cm (A,B); 1 cm (C). <a href="https://www.jove.com/files/ftp_upload/64747/64747fig03large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/64747/64747fig04.jpg" /> 
Figure 4: Holder designed for sharpening the cutting edge of increment corers. (A)&#160;Teflon block to guide and stabilize the increment corer. (B) Teflon block to fix the starter of the cordless drill. (C) Hexagon wrench to fix A at other positions on the board, depending on the type of corer used. (D) Clamp to fix the cordless drill. (E) Placing the conical grinding stone inside the cutting edge. (F) Placing the straight grinding stone outside to deburr the cutting edge. <a href="https://www.jove.com/files/ftp_upload/64747/64747fig04large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/64747/64747fig05.jpg" /> 
Figure 5: Application of the water-soluble tape. (A) Cut the tape to the length needed to cover the core surface. (B) Place the tape on the prepared surface of the core. (C) Cut the section by holding the edge of the tape in one hand. (D) Place the tape with the section facing down on a glass slide and add water to separate the tape from the section. <a href="https://www.jove.com/files/ftp_upload/64747/64747fig05large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 6" class="xfigimg" src="/files/ftp_upload/64747/64747fig06.jpg" /> 
Figure 6: Micro section of a conifer showing a &#34;blue ring&#34;. Latewood cell walls not lignified and, for this, blue, in the ring of the year 1974 (magnified above the slide showing the long section). Scale bar = 1 cm. <a href="https://www.jove.com/files/ftp_upload/64747/64747fig06large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Watch this Scientific Journal Video about Advanced Workflow for Taking High-Quality Increment Cores - New Techniques and Devices at JoVE.com

Advanced Workflow for Taking High-Quality Increment Cores - New Techniques and Devices

Here, we present a protocol on how to avoid micro cracks in increment cores by applying a cordless drill with a torque multiplier to minimize problems when coring trees, as well as its effect on preparing long micro sections. This protocol also includes a procedure to sharpen corers in the field.

In dendroecological research, precise dating of each single growth ring is a basic requirement for all studies, focusing on ring-width variations only, ...

advanced-workflow-for-taking-high-quality-increment-cores-new

Research

JoVE Journal

Environment

1.9K Views.  Swiss Federal Research Institute WSL. Here, we present a protocol on how to avoid micro cracks in increment cores by applying a cordless drill with a torque multiplier to minimize problems when coring trees, as well as its effect on preparing long micro sections. This protocol also includes a procedure to sharpen corers in the field.

Video: Advanced Workflow for Taking High-Quality Increment Cores - New Techniques and Devices

Methods for Performing Crosses in Setaria viridis, a New Model System for the Grasses

Setaria viridis is an emerging model system for C4 grasses. It is closely related to the bioenergy feed stock switchgrass and the grain crop foxtail millet. Recently, the 510 Mb genome of foxtail millet, S. italica, has been sequenced 1,2 and a 25x coverage genome sequence of the weedy relative S. viridis is in progress. S. viridis has a number of characteristics that make it a potentially excellent model genetic system including a rapid generation time, small stature, simple growth requirements, prolific seed production 3 and developed systems for both transient and stable transformation 4. However, the genetics of S. viridis is largely unexplored, in part, due to the lack of detailed methods for performing crosses. To date, no standard protocol has been adopted that will permit rapid production of seeds from controlled crosses.
The protocol presented here is optimized for performing genetic crosses in S. viridis, accession A10.1. We have employed a simple heat treatment with warm water for emasculation after pruning the panicle to retain 20-30 florets and labeling of flowers to eliminate seeds resulting from newly developed flowers after emasculation. After testing a series of heat treatments at permissive temperatures and varying the duration of dipping, we have established an optimum temperature and time range of 48 &deg;C for 3-6 min. By using this method, a minimum of 15 crosses can be performed by a single worker per day and an average of 3-5 outcross progeny per panicle can be recovered. Therefore, an average of 45-75 outcross progeny can be produced by one person in a single day. Broad implementation of this technique will facilitate the development of recombinant inbred line populations of S. viridis X S. viridis or S. viridis X S. italica, mapping mutations through bulk segregant analysis and creating higher order mutants for genetic analysis.

Methods for Performing Crosses in Setaria viridis, a New Model System for the Grasses

We have developed a methodology for performing crosses in Setaria viridis (S. viridis). The method involves pruning the panicle prior to a hot water treatment to kill viable pollen. Crosses are performed following a well-controlled growth regime and typically result in the recovery of 1 to 7 cross-pollinated seed/s per panicle.

Setaria viridis is an emerging model system for C4 grasses. It is closely related to the bioenergy feed stock switchgrass and the grain crop foxtail ...

Surface Renewal: An Advanced Micrometeorological Method for Measuring and Processing Field-Scale Energy Flux Density Data

Advanced micrometeorological methods have become increasingly important in soil, crop, and environmental sciences. For many scientists without formal training in atmospheric science, these techniques are relatively inaccessible. Surface renewal and other flux measurement methods require an understanding of boundary layer meteorology and extensive training in instrumentation and multiple data management programs. To improve accessibility of these techniques, we describe the underlying theory of surface renewal measurements, demonstrate how to set up a field station for surface renewal with eddy covariance calibration, and utilize our open-source turnkey data logger program to perform flux data acquisition and processing. The new turnkey program returns to the user a simple data table with the corrected fluxes and quality control parameters, and eliminates the need for researchers to shuttle between multiple processing programs to obtain the final flux data. An example of data generated from these measurements demonstrates how crop water use is measured with this technique. The output information is useful to growers for making irrigation decisions in a variety of agricultural ecosystems. These stations are currently deployed in numerous field experiments by researchers in our group and the California Department of Water Resources in the following crops: rice, wine and raisin grape vineyards, alfalfa, almond, walnut, peach, lemon, avocado, and corn.

Surface renewal is a micrometeorological method that is being used increasingly to determine energy fluxes, but its technical complexity makes it inaccessible to a broad audience. We describe the steps needed to set up and calibrate a surface renewal field station, to acquire and process data, and to correctly interpret results.

Advanced micrometeorological methods have become  increasingly important in soil, crop, and environmental sciences.  For many scientists without formal ...

Empirical, Metagenomic, and Computational Techniques Illuminate the Mechanisms by which Fungicides Compromise Bee Health

Growers often use fungicide sprays during bloom to protect crops against disease, which exposes bees to fungicide residues. Although considered &#34;bee-safe,&#34; there is mounting evidence that fungicide residues in pollen are associated with bee declines (for both honey and bumble bee species). While the mechanisms remain relatively unknown, researchers have speculated that bee-microbe symbioses are involved. Microbes play a pivotal role in the preservation and/or processing of pollen, which serves as nutrition for larval bees. By altering the microbial community, it is likely that fungicides disrupt these microbe-mediated services, and thereby compromise bee health. This manuscript describes the protocols used to investigate the indirect mechanism(s) by which fungicides may be causing colony decline. Cage experiments exposing bees to fungicide-treated flowers have already provided the first evidence that fungicides cause profound colony losses in a native bumble bee (Bombus impatiens). Using field-relevant doses of fungicides, a series of experiments have been developed to provide a finer description of microbial community dynamics of fungicide-exposed pollen. Shifts in the structural composition of fungal and bacterial assemblages within the pollen microbiome are investigated by next-generation sequencing and metagenomic analysis. Experiments developed herein have been designed to provide a mechanistic understanding of how fungicides affect the microbiome of pollen-provisions. Ultimately, these findings should shed light on the indirect pathway through which fungicides may be causing colony declines.

Microbial consortia within bumble bee hives enrich and preserve pollen for bee larvae. Using next generation sequencing, along with laboratory and field-based experiments, this manuscript describes protocols used to test the hypothesis that fungicide residues alter the pollen microbiome, and colony demographics, ultimately leading to colony loss.

Growers often use fungicide sprays during bloom to protect crops against disease, which exposes bees to fungicide residues. Although considered ...

Techniques for Investigating the Anatomy of the Ant Visual System

This article outlines a suite of techniques in light microscopy (LM) and electron microscopy (EM) which can be used to study the internal and external eye anatomy of insects. These include traditional histological techniques optimized for work on ant eyes and adapted to work in concert with other techniques such as transmission electron microscopy (TEM) and scanning electron microscopy (SEM). These techniques, although vastly useful, can be difficult for the novice microscopist, so great emphasis has been placed in this article on troubleshooting and optimization for different specimens. We provide information on imaging techniques for the entire specimen (photo-microscopy and SEM) and discuss their advantages and disadvantages. We highlight the technique used in determining lens diameters for the entire eye and discuss new techniques for improvement. Lastly, we discuss techniques involved in preparing samples for LM and TEM, sectioning, staining, and imaging these samples. We discuss the hurdles that one might come across when preparing samples and how best to navigate around them.

This article outlines a suite of techniques in light and electron microscopy to study the internal and external eye anatomy of insects. These include several traditional techniques optimized for work on ant eyes, detailed troubleshooting, and suggestions for optimization for different specimens and regions of interest.

This article outlines a suite of techniques in light microscopy (LM) and electron microscopy (EM) which can be used to study the internal and external eye ...

Multimodal Hierarchical Imaging of Serial Sections for Finding Specific Cellular Targets within Large Volumes

Targeting specific cells at ultrastructural resolution within a mixed cell population or a tissue can be achieved by hierarchical imaging using a combination of light and electron microscopy. Samples embedded in resin are sectioned into arrays consisting of ribbons of hundreds of ultrathin sections and deposited on pieces of silicon wafer or conductively coated coverslips. Arrays are imaged at low resolution using a digital consumer like smartphone camera or light microscope (LM) for a rapid large area overview, or a wide field fluorescence microscope (fluorescence light microscopy (FLM)) after labeling with fluorophores. After post-staining with heavy metals, arrays are imaged in a scanning electron microscope (SEM). Selection of targets is possible from 3D reconstructions generated by FLM or from 3D reconstructions made from the SEM image stacks at intermediate resolution if no fluorescent markers are available. For ultrastructural analysis, selected targets are finally recorded in the SEM at high-resolution (a few nanometer image pixels). A ribbon-handling tool that can be retrofitted to any ultramicrotome is demonstrated. It helps with array production and substrate removal from the sectioning knife boat. A software platform that allows automated imaging of arrays in the SEM is discussed. Compared to other methods generating large volume EM data, such as serial block-face SEM (SBF-SEM) or focused ion beam SEM (FIB-SEM), this approach has two major advantages: (1) The resin-embedded sample is conserved, albeit in a sliced-up version. It can be stained in different ways and imaged with different resolutions. (2) As the sections can be post-stained, it is not necessary to use samples strongly block-stained with heavy metals to introduce contrast for SEM imaging or render the tissue blocks conductive. This makes the method applicable to a wide variety of materials and biological questions. Particularly prefixed materials e.g., from biopsy banks and pathology labs, can directly be embedded and reconstructed in 3D.

This protocol targets specific cells in tissue for imaging at nanoscale resolution using a scanning electron microscope (SEM). Large numbers of serial sections from resin-embedded biological material are first imaged in a light microscope to identify the target and then in a hierarchical manner in the SEM.

Targeting specific cells at ultrastructural resolution within a mixed cell population or a tissue can be achieved by hierarchical imaging using a ...

Developmental Biology

High-Throughput Tree Ring Analysis and Density Estimation

Tree Core Analysis with X-ray Computed Tomography

An X-ray computed tomography (CT) toolchain is presented to obtain tree-ring width (TRW), maximum latewood density (MXD), other density parameters, and quantitative wood anatomy (QWA) data without the need for labor-intensive surface treatment or any physical sample preparation. The focus here is on increment cores and scanning procedures at resolutions ranging from 60 &#181;m down to 4 &#181;m. Three scales are defined at which wood should be looked at: (i) inter-ring scale, (ii) ring scale, i.e., tree-ring analysis and densitometry scale, as well as (iii) anatomical scale, the latter approaching the conventional thin-section quality. Custom-designed sample holders for each of these scales enable high-throughput scanning of multiple increment cores. A series of software routines were specifically developed to efficiently treat three-dimensional X-ray CT images of the tree cores for TRW and densitometry. This work briefly explains the basic principles of CT, which are needed for a proper understanding of the protocol. The protocol is presented for some known species that are commonly used in dendrochronology. The combination of rough density estimates, TRW and MXD data, as well as quantitative anatomy data, allows us to broaden and deepen current analyses for climate reconstructions or tree response, as well as further develop the field of dendroecology/climatology and archeology.

Author Spotlight: Advancements in X-ray CT Tool Chain for Tree Core Analysis

Here we show how to process tree cores with an X-ray computed tomography toolchain. Except for chemical extraction for some purposes, no further physical lab treatment is needed. The toolchain can be used for biomass estimations, for obtaining MXD/tree-ring width data as well as for obtaining quantitative wood anatomy data.

An X-ray computed tomography (CT) toolchain is presented to obtain tree-ring width (TRW), maximum latewood density (MXD), other density parameters, and ...

Protocol for Field to Lab Tree Ring Digitization in Dendroecological Studies

A New Workflow for Sampling and Digitizing Increment Cores

Here we present a new workflow from taking increment cores in the field, storing and transporting them to the lab, to digitizing their tree rings for further analyses for subsequent dendroecological analyses. The procedure involves the use of new sample carriers for increment cores. These new G&#228;rtner Schneider Core (GSC) holders are designed using three-dimensional (3D) modeling software and finally printed with a 3D printer. Using these mounts from the beginning in the field, the cores can directly be cut with a core microtome, and their surface can then be digitized without further rearrangement using a new high-resolution image-capturing system. They are thus available for direct analysis. This system allows digitizing tree rings from cores and disks, and also taking images from long micro sections (up to 40 cm) using transmitted light. This feature is of special interest for dendroecological and geomorphic applications to identify the onset of any disturbance in micro sections cut with a core-microtome.

Author Spotlight: Innovative Device Development for Advancing Dendroecology and Wood Anatomy Research

We present a protocol to use 3D-printed mounts to fix increment cores in the field without the necessity of unpacking and gluing them on wooden mounts. The new GSC holder allows placing the cores in a core microtome to cut their surface and directly transfer them to digital image capturing.

Here we present a new workflow from taking increment cores in the field, storing and transporting them to the lab, to digitizing their tree rings for ...

Parallel Active Cantilever Arrays in AFMS to Enable High-Throughput Inspections

Active Probe Atomic Force Microscopy with Quattro-Parallel Cantilever Arrays for High-Throughput Large-Scale Sample Inspection

An Atomic Force Microscope&#160;(AFM) is&#160;a powerful and versatile tool for nanoscale surface studies to capture 3D topography images of samples. However, due to their limited imaging throughput, AFMs have not been widely adopted for large-scale inspection purposes. Researchers have developed high-speed AFM systems to record dynamic process videos in chemical and biological reactions at tens of frames per second, at the cost of a small imaging area of up to several square micrometers. In contrast, inspecting large-scale nanofabricated structures, such as semiconductor wafers, requires nanoscale spatial resolution imaging of a static sample over hundreds of square centimeters with high productivity. Conventional AFMs use a single passive cantilever probe with an optical beam deflection system, which can only collect one pixel at a time during AFM imaging, resulting in low imaging throughput. This work utilizes an array of active cantilevers with embedded piezoresistive sensors and thermomechanical actuators, which allows simultaneous multi-cantilever operation in parallel operation for increased imaging throughput. When combined with large-range nano-positioners and proper control algorithms, each cantilever can be individually controlled to capture multiple AFM images. With data-driven post-processing algorithms, the images can be stitched together, and defect detection can be performed by comparing them to the desired geometry. This paper introduces principles of the custom AFM using the active cantilever arrays, followed by a discussion on practical experiment considerations for inspection applications. Selected example images of silicon calibration grating, highly-oriented pyrolytic graphite, and extreme ultraviolet lithography masks are captured using an array of four active cantilevers (&#34;Quattro&#34;) with a 125 &#181;m tip separation distance. With more engineering integration, this high-throughput, large-scale imaging tool can provide 3D metrological data for extreme ultraviolet (EUV) masks, chemical mechanical planarization (CMP) inspection, failure analysis, displays, thin-film step measurements, roughness measurement dies, and laser-engraved dry gas seal grooves.

Author Spotlight: Introduction to Active Probe Atomic Force Microscopy with Quattro-Parallel Cantilever Arrays

Large-scale sample inspection with nanoscale resolution has a wide range of applications, especially for nanofabricated semiconductor wafers. Atomic force microscopes can be a great tool for this purpose, but are limited by their imaging speed. This work utilizes parallel active cantilever arrays in AFMs to enable high-throughput and large-scale inspections.

An Atomic Force Microscope&#160;(AFM) is&#160;a powerful and versatile tool for nanoscale surface studies to capture 3D topography images of samples. ...

Engineering

A Sectioning, Coring, and Image Processing Guide for High-Throughput Cortical Bone Sample Procurement and Analysis for Synchrotron Micro-CT

Bone is a dynamic and mechanically active tissue that changes in structure over the human lifespan. The products of the bone remodeling process have been studied substantially using traditional two-dimensional techniques. Recent advancements in X-ray imaging technology via desktop micro-computed tomography (&#181;CT) and synchrotron radiation micro-computed tomography (SR&#181;CT) have allowed for the acquisition of high-resolution three-dimensional (3D) scans of a larger field of view (FOV) than other 3D imaging techniques (e.g., SEM) providing a more complete picture of microscopic structures within human cortical bone. The specimen should be accurately centered within the FOV, however, to limit the appearance of streak artifacts known to impact data analysis. Previous studies have reported procurement of irregularly shaped rectilinear bone blocks that result in imaging artifacts due to uneven edges or image truncation. We have applied a geological sampling protocol (coring) to procure consistently sized cortical bone core specimens for SR&#181;CT experiments from the anterior aspect of human femora. This coring method is efficient and minimally destructive to tissue. It creates uniform cylindrical samples that decrease imaging artifacts by nature of being isometric during rotation and providing a uniform path length for X-ray beams throughout scanning. Image processing of X-ray tomographic data of cored and irregularly shaped samples confirms the potential of the technique to improve visualization and analysis of cortical bone microarchitecture. A goal of this protocol is to deliver a reliable and repeatable method for the extraction of cortical bone cores that is adaptable for various types of high-resolution bone imaging experiments. An overarching goal of the work is to create a standardized cortical bone procurement for SR&#181;CT that is affordable, consistent, and straightforward. This procedure may further be adapted by researchers in related fields who commonly evaluate hard composite materials such as in biological anthropology, geosciences, or material sciences.

We employed a geological (coring) sampling protocol to procure cortical bone specimens of uniform size for SR&#181;CT experiments from the anterior aspect of human femora. This method is minimally destructive, efficient, results in cylindrical specimens that minimize imaging artifacts from irregular sample shapes and improves microarchitectural visualization and analysis.

Bone is a dynamic and mechanically active tissue that changes in structure over the human lifespan. The products of the bone remodeling process have been ...

Biology

Array Tomography Workflow for the Targeted Acquisition of Volume Information using Scanning Electron Microscopy

Electron microscopy is applied in biology and medicine for imaging of cellular and structural details at nanometer resolution. Historically, Transmission Electron Microscopy (TEM) provided insight into cell ultrastructure, but in the recent decade, the development of modern Scanning Electron Microscopes (SEM) has changed the way of looking inside the cells. Even though the resolution of TEM is superior when protein-level structural details are needed, SEM-resolution is sufficient for the majority of the organelle-level cell biology-related questions. The advancement in technology enabled automatic volume acquisition solutions such as Serial block-face imaging (SBF-SEM) and Focused ion beam SEM (FIB-SEM). Nevertheless, to this day, these methods remain inefficient when the identification and navigation to areas of interest are crucial. Without the means for precise localization of target areas before imaging, operators need to acquire much more data than they need (in SBF-SEM), or, even worse, prepare many grids and image them all (in TEM). We propose the strategy of "lateral screening" using Array Tomography in SEM, which facilitates the localization of areas of interest, followed by automated imaging of the relevant fraction of the total sample volume. Array tomography samples are conserved during imaging, and they can be arranged into section libraries ready for repeated imaging. Several examples are shown in which lateral screening enables us to analyze structural details that are incredibly challenging to access with any other method.

We describe the preparation of ribbons of serial sections and their collection on large transfer support for use as Array Tomography samples, along with automated imaging procedures in a scanning electron microscope. The protocol allows screening, retrieval, and targeted imaging of local, rare events, and the acquisition of large data volumes.

Electron microscopy is applied in biology and medicine for imaging of cellular and structural details at nanometer resolution. Historically, Transmission ...

Advanced Workflow for Taking High-Quality Increment Cores - New Techniques and Devices

Podsumowanie

Przeglądaj więcej filmów

Rozdziały w tym wideo

Methods for Performing Crosses in Setaria viridis, a New Model System for the Grasses

Surface Renewal: An Advanced Micrometeorological Method for Measuring and Processing Field-Scale Energy Flux Density Data

Empirical, Metagenomic, and Computational Techniques Illuminate the Mechanisms by which Fungicides Compromise Bee Health

Techniques for Investigating the Anatomy of the Ant Visual System

Multimodal Hierarchical Imaging of Serial Sections for Finding Specific Cellular Targets within Large Volumes

Tree Core Analysis with X-ray Computed Tomography

A New Workflow for Sampling and Digitizing Increment Cores

Active Probe Atomic Force Microscopy with Quattro-Parallel Cantilever Arrays for High-Throughput Large-Scale Sample Inspection

A Sectioning, Coring, and Image Processing Guide for High-Throughput Cortical Bone Sample Procurement and Analysis for Synchrotron Micro-CT

Array Tomography Workflow for the Targeted Acquisition of Volume Information using Scanning Electron Microscopy