Name: analysis of n-glycans from raphanus sativus cultivars using pngase h+
Uploaded: 2018-06-25T15:00:00
Description: Watch this Scientific Journal Video about Analysis of N-glycans from Raphanus sativus Cultivars Using PNGase H+ at JoVE.com

Este trabalho foi financiado em parte pela Fundação ciência Natural da China (conceder números 31471703, A0201300537 e 31671854, J.V. e ll, conceder número 31470435 para G.Y.) e o plano de talentos estrangeiros 100 (número de concessão JSB2014012 de J.V.).

1. coleta de amostra
<ol><li>Compra de diferentes cultivares de fresco rabanete (Raphanus sativus L.).</li>
</ol>2. isolamento da proteína de rabanete
<ol><li>Homogeneizar a cerca de 100 g de rabanete fresco com um misturador de cozinha para 10 min.</li>
	<li>Transferi o chorume para um tubo de centrífuga de 50 mL e centrifugação a 14.000 × g a 4 ° C por 20 min remover o material insolúvel.</li>
	<li>Transferir o sobrenadante cuidadosamente para um novo tubo de centrif...

<ol>
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J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Strategies+for+the+Structural+Analysis+of+Carbohydrates.">Strategies for the Structural Analysis of Carbohydrates.</a> Comprehensive Glycoscience. , 1-68 (2007).</li><li>Huang, Y., Mechref, Y., Novotny, M. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microscale+nonreductive+release+of+O-linked+glycans+for+subsequent+analysis+through+MALDI+mass+spectrometry+and+capillary+electrophoresis.">Microscale nonreductive release of O-linked glycans for subsequent analysis through MALDI mass spectrometry and capillary electrophoresis.</a> Analytical Chemistry. 73 (24), 6063-6069 (2001).</li><li>Triguero, A., 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=Chemical+and+enzymatic+N-glycan+release+comparison+for+N-glycan+profiling+of+monoclonal+antibodies+expressed+in+plants.">Chemical and enzymatic N-glycan release comparison for N-glycan profiling of monoclonal antibodies expressed in plants.</a> Analytical Biochemistry. 400 (2), 173-183 (2010).</li><li>Tretter, V., Altmann, F., Marz, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Peptide-N4-(N-acetyl-&#946;-glucosaminyl)asparagine+amidase+F+cannot+release+glycans+with+fucose+attached+&#945;1+&#8594;+3+to+the+asparagine-linked+N-acetylglucosamine+residue.">Peptide-N4-(N-acetyl-&#946;-glucosaminyl)asparagine amidase F cannot release glycans with fucose attached &#945;1 &#8594; 3 to the asparagine-linked N-acetylglucosamine residue.</a> European Journal of Biochemistry. 199 (3), 647-652 (1991).</li><li>Fan, J. -. 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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=Rapid+sample+preparation+methodology+for+plant+N-.glycan+analysis+using+acid-stable+PNGase+H+.">Rapid sample preparation methodology for plant N-.glycan analysis using acid-stable PNGase H+.</a> Journal of Agricultural and Food Chemistry. 63 (48), 10550-10555 (2015).</li><li>Wang, T., Hu, X. -. C., Cai, Z. -. P., Voglmeir, J., Liu, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Qualitative+and+Quantitative+Analysis+of+Carbohydrate+Modification+on+Glycoproteins+from+seeds+of+Ginkgo+biloba.">Qualitative and Quantitative Analysis of Carbohydrate Modification on Glycoproteins from seeds of Ginkgo biloba.</a> Journal of Agricultural and Food Chemistry. 65 (35), 7669-7679 (2017).</li><li>Ceroni, A., 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=GlycoWorkbench:+a+tool+for+the+computer-assisted+annotation+of+mass+spectra+of+glycans.">GlycoWorkbench: a tool for the computer-assisted annotation of mass spectra of glycans.</a> Journal of Proteome Research. 7 (4), 1650-1659 (2008).</li></ol>

Apresentamos aqui o protocolo permite a comparação dos perfis de vários cultivares de rabanete - glicano N. Uma vantagem significativa do método comparado com protocolos existentes é que nenhuma alteração de tampão entre a liberação enzimática de N- os glicanos e a reação de derivatização com 2-AB é necessárias. O passo mais crítico deste procedimento é a purificação do N- os glicanos usando a coluna SPE, como falha para remover os sais ou outras impurezas na mistura de rea...

Os N- glicanos em plantas têm chamado a atenção crescente nos últimos anos, como pesquisas anteriores destacou os N- glicanos como uma fonte potencial de reacções imunológicas que podem provocar respostas alérgicas1,2 . Foi demonstrado anteriormente que os N- glicanos em glicoproteínas de planta pode afetar a atividade catalítica de3,4, estabilidade térmica e dobramento5,6 ou Localização subcellular e secreção7. A fim de correlacionar as estruturas glicano com suas respectivas funções, os N- glicanos deve ser lançados de glicoproteínas ou quimicamente ou enzimaticamente. O método químico clássico para liberar N- e O- os glicanos é β-eliminação, em que o tratamento alcalino amostra é acompanhado por redução com borohidreto para produzir um alditol8. No entanto, este procedimento impede a rotulagem com um fluoróforo e causa deterioração significativa de unidades de monossacarídeo do redutor final da estrutura de glicano. Deglycosylation química, com base no tratamento de hidróxido de amônio/carbonato é também um método alternativo comumente usado9. Nenhum desses métodos de liberação química degrada proteínas intactas, que permite a análise de espectrometria de massa de glicano sem rótulo piscinas sem a interferência de fragmentos de peptídeo na mesma faixa de massa. No entanto, um inconveniente desses métodos é a taxa de aumento da degradação de α1, 3-fucosilado N- os glicanos, uma estrutura comum de carboidrato encontrado em plantas,10. Como alternativa, os métodos de liberação enzimática usando peptídeo:N- glycanases (PNGases, CE 3.5.1.52) são também amplamente aplicados. Recombinante PNGase F (de Flavobacterium meningosepticum) é a escolha mais comum e permite a libertação de todos os tipos de N- os glicanos, exceto as estruturas portadoras de um núcleo α1, fucose 311,12. Portanto, A PNGase (isoladas de sementes amêndoas) geralmente é usada para a análise da planta Nos glicanos -13. No entanto, esta enzima deglycosylates apenas proteoliticamente-derivado glicopeptídeos e é incapaz de deglycosylate nativo glicoproteínas14. Portanto, um exame de amostra de várias etapas é necessário antes de mais análise, que provoca a perda extensa dos glicanos, especialmente aqueles de baixa abundância15. O objetivo geral do método é apresentar um fluxo de trabalho otimizado para N- glicano liberação e rotulagem de forma simples e robusta de fluorescência. A lógica subjacente é que o PNGase H+, que recentemente foi descoberto em Terriglobus roseus e pode ser expressa de recombinantes em e. coli, pode hidrolisar os N- glicanos diretamente do andaime proteína em ácido condições de16. A principal vantagem de usar PNGase H+ sobre métodos alternativos é que reações de rotulagem fluorescentes podem ser executadas em tubo de reação mesmo sem alterar a reação amortecedores17,18. As condições de preparação simples e alta recuperação de oligossacarídeos de baixo-abundância fazem esse método uma ferramenta valiosa na análise do N- os glicanos. Este protocolo é apropriado para a análise de N- os glicanos de várias espécies de plantas.

<table border="0" cellpadding="0" cellspacing="0" style="width:880px;" width="880">
	<colgroup>
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		<col />
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	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:347px;">Chemicals:</td>
			<td style="width:123px;"></td>
			<td style="width:207px;"></td>
			<td style="width:204px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Trichloroacetic acid&#160;</td>
			<td>SCR, Shanghai</td>
			<td>80132618</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Acetic acid glacial</td>
			<td>Huada, Guangzhou</td>
			<td>64-19-7</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Acetonitrile</td>
			<td>General-reagent</td>
			<td>G80988C</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Trifluoroacetic acid</td>
			<td>Energy chemical</td>
			<td>W810031</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">2-aminobenzamide</td>
			<td>Heowns, Tianjin</td>
			<td>A41900</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Sodium cyanoborohydride</td>
			<td>J&#38;K Scientific Ltd</td>
			<td>314162</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Dimethyl sulfoxide</td>
			<td>Huada, Guangzhou</td>
			<td>67-68-5</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">2AB-labeled dextran ladder, 200 pmol</td>
			<td>Agilent Technologies</td>
			<td>AT-5190-6998</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">6-Aza-2-thiothymine&#160;</td>
			<td>Sigma</td>
			<td>275514</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tools/Materials:</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Kitchen blender</td>
			<td>Bear, Guangzhou</td>
			<td>LLJ-A10T1</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Centrifuge</td>
			<td>Techcomp</td>
			<td>CT15RT</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Centrifugal Evaporator</td>
			<td>Hualida, Taicang</td>
			<td>LNG-T120</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">SPE column</td>
			<td>Supelco</td>
			<td>Supelclean ENVI Carb SPE column</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">MALDI-TOF mass spectrometer</td>
			<td>Bruker</td>
			<td>Autoflex</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">HPLC Analysis:</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">High-recovery HPLC vial</td>
			<td>Agilent Technologies</td>
			<td>&#160;# 5188-2788</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">HPLC System</td>
			<td>Shimadzu</td>
			<td>Nexera</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Fluorescence Detector for HPLC</td>
			<td>Shimadzu</td>
			<td>RF-20Axs&#160;</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Column oven</td>
			<td>Hengxin</td>
			<td>CO-2000</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">HPLC Column</td>
			<td>Waters</td>
			<td>Acquity UPLC BEH glycan column</td>
			<td>2.1 &#215; 150 mm, 1.7 &#956;m particle size</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">LCMS-grade Water</td>
			<td>Merck Millipore</td>
			<td>#WX00011</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">LCMS-grade Acetonitrile</td>
			<td>Merck Millipore</td>
			<td># 100029</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Formic acid</td>
			<td>Aladdin</td>
			<td>F112034</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Ammonia solution</td>
			<td>Aladdin</td>
			<td>A112080</td>
			<td></td>
		</tr>
	</tbody>
</table>

analysis of n-glycans from raphanus sativus cultivars using pngase h+

Nos últimos anos, as metades de hidrato de carbono de plantas têm recebido atenção considerável, como eles são uma fonte potencial de cross-reactive, alergia, provocando reacções imunológicas. Além disso, estruturas de carboidratos também desempenham um papel fundamental no metabolismo da planta. Aqui, apresentamos um método simples e rápido para preparar e analisar N- os glicanos de diferentes cultivares de rabanete (Raphanus sativus) usando um N- glycanase específico para o lançamento de estruturas de carboidratos de origem vegetal. Para conseguir isso, cru ácido tricloroacético precipitados de rabanete homogenates foram tratados com PNGase H+e rotulados usando 2-aminobenzamide como uma etiqueta fluorescente. N- glicano amostras etiquetadas foram analisadas posteriormente pela separação de ultra desempenho cromatografia líquida (UPLC) e laser assistida por matriz dessorção ionização tempo de voo (MALDI-TOF) espectrometria de massa para uma detalhada estrutural avaliação e a quantificação relativas abundancies das estruturas rabanete-derivados N- glicano. Este protocolo também pode ser usado para a análise de N- os glicanos de várias outras espécies de plantas e pode ser útil para posterior investigação da função e efeitos da N- os glicanos na saúde humana.

A Figura 1 mostra uma visão esquemática do protocolo descrito, incluindo o isolamento de proteínas (Glico-) de rabanete, a preparação de N- os glicanos, a análise UPLC e a análise de MALDI-TOF-MS desses componentes. A Figura 2 mostra representativas cromatogramas UPLC do pyrazole N- os glicanos dos cultivars rabanete analisados. A Figura 3 mostra os resultados obtidos das est...

Watch this Scientific Journal Video about Analysis of N-glycans from Raphanus sativus Cultivars Using PNGase H+ at JoVE.com

Analysis of N-glycans from Raphanus sativus Cultivars Using PNGase H+

Análise dos N- glicanos de Raphanus sativus cultivares usando PNGase H+

Nós descrevemos um método simples e rápido para a preparação e análise de N- os glicanos de diferentes cultivares de rabanete (Raphanus sativus).

In recent years, the carbohydrate moieties of plants have received considerable attention, as they are a potential source of cross-reactive, ...

This method can help answer questions in the field of plant Glycoscience such as how N-glycans of plant origin can be quickly analyzed and characterized. The main advantage of this technique is that it doesn't require any buffer changes between the enzymatic release of N-glycans and the derivatization reactions with 2-aminobenzamide. Demonstrating the procedure will be miss Ya-Min Du, and miss Shen-Li Zhen;two graduate students from the Glycomics and Glycan Bioengineering Research Center at Nanjing Agricultural University.To begin isolating the proteins, use a kitchen blender to homogenize approximately 100 grams of fresh radish for ten minutes. Transfer the resulting slurry to a 50 milliliter centrifuge tube. Centrifuge at 14, 000 times gravity at four degrees Celsius for 20 minutes to remove the insoluble material.After this, carefully transfer the supernatant into a new 50 milliliter centrifuge tube. Add an equal amount of a two molar trichloroacetic acid solution. Centrifuge at 14, 000 times gravity at four degrees Celsius for 30 minutes.And then remove the supernatant. Wash the pellet with 20 milliliters of deionized water. Centrifuge again at 14, 000 times gravity at four degrees Celsius for five minutes to remove soluble oligosaccharides and polysaccharides.Then, re-suspend the pellet with one milliliter of deionized water. And transfer the protein solution to a fresh 1.5 milliliter centrifuge tube. To begin preparing the N-glycans, mix 50 microliters of the protein solution with 0.2 milliunits of recombinant PNGase H+in ten millimolar acetic acid.Incubate at 37 degrees Celsius for 12 hours. Then, centrifuge at 14, 000 times gravity for five minutes to remove the extra protein and enzyme. Transfer the supernatant to a fresh 1.5 milliliter centrifuge tube until ready to purify the N-glycans.To begin purifying the N-glycans, wash the solid phase extraction column with three milliliters of water. Activate the column by adding three milliliters of an 80 percent acetonitrile solution containing trifluoroacetic acid. Then, three milliliters of deionized water to equilibrate the column.Transfer the sample onto the column and discard the flow through. Wash the column by adding 1.5 milliliters of deionized water. Failure to remove salts and add impurities in the sample mixture may negatively impact the efficiency of the fluorescence derivatization reaction.Then, add 1.5 milliliters of an aqueous 20 percent acetonitrile solution containing 0.1 percent TFA to elute the N-glycans into a two milliliter tube. Remove the solvent by centrifugal evaporation at room temperature, evaporating until the sample is completely dry. To begin fluorescence derivatization of N-glycans, prepare the 2-AB solution as outlined in the text protocol.Add five microliters of the prepared 2-AB solution to the dried samples. Vortex until each sample is completely dissolved. Incubate at 65 degrees Celsius for two hours.After this, let the sample cool to room temperature for five minutes. Add five microliters of deionized water and 40 microliters of acetonitrile to the cooled sample. Centrifuge at 14, 000 times gravity for three minutes.Then, transfer 48 microliters of the resulting supernatant into a 300 microliter high recovery HPLC vial. Store at negative 20 degrees Celsius for up to one month or when ready to use. To begin profiling, use a standard UPLC system equipped with a hydrophobic interaction liquid chromatography UPLC glycan column and connect it to an online fluorescence detector to analyze the samples.Set the excitation and emission wavelengths to 330 nanometers and 420 nanometers, respectively. Set the column temperature to 60 degrees Celsius. Next, inject 45 microliters of the sample into the UPLC system.Elute the N-glycans by UPLC using retention times of 15 and 40 minutes. Using two milliliters centrifuge tubes collect each observed UPLC peak fraction. Then, use centrifugal evaporation to dry the samples.Inject approximately one picomole of 2-AB labeled dextran standard to calibrate the plant N-glycan profiling and to assign their elution times and to standardize glucose units. To begin MALDI-TOF analysis, dissolve the dried samples in five microliters of LCMS grade water. Mix one microliter of this solution with one microliter of ATT solution.Then, pipet this mixture into the MALDI-TOF sample carrier. After this, use a MALDI-TOF mass spectrometer instrument to analyze the samples as outlined in the text protocol. In this study, N-glycan profiles are compared from various cultivars of radish.Representative UPLC chromatograms show that derivatized N-glycans and the relative abundance of each and different radishes can be successfully analyzed through this process. After this, MALDI-TOF mass spectroscopy is used to analyze the structures of 2-AB derivatized N-glycans. Though this method can provide insight into the composition of N-glycans from different Raphanus sativus cultivars, it can also be applied to other systems such as peanuts or other allergenic plant species.Once mastered, samples can be prepared within two working days which already includes the UPLC analysis procedure. When attempting this procedure, it's important to remember that the solid phase extraction columns used to isolate the N-glycans are sufficiently moistened with distilled water prior to sample application. Otherwise, binding of N-glycans to the column may be hampered.For additional verification of N-glycan structures, other methods can also be used. This could be, for example, sequential exoglycosidase treatments or MALDI-TOF MS N based fragmentation techniques. After its development, this technique paved the way for researchers in the field of Glycoscience to explore the composition and function of plant N-glycans more rapidly.After watching this video, you should have a good understanding of how to prepare N-glycans from different cultivars of radish using PNGase H+and how to analyze the 2-aminobenzamide derivatives by UPLC. Don't forget that working with trifluoroacetic acid and trichloroacetic acid can be extremely hazardous and precautions such as wearing protective gear should always be taken while performing this procedure.

Research

JoVE Journal

Biochemistry

Analysis of N-glycans from Raphanus sativus Cultivars Using PNGase H+

High-resolution Single Particle Analysis from Electron Cryo-microscopy Images Using SPHIRE

Análise de uma única partícula de alta resolução a partir de imagens de crio-microscopia de elétrons usando SPHIRE

SPHIRE (SPARX for High-Resolution Electron Microscopy) is a novel open-source, user-friendly software suite for the semi-automated processing of single ...

Bioquímica

Isolation of Intermediate Filament Proteins from Multiple Mouse Tissues to Study Aging-associated Post-translational Modifications

Isolamento de Proteínas de Filamento Intermediário a partir de Tecidos Múltiplos de Rato para Estudar Modificações Pós-Translacionais associadas ao Envelhecimento

Intermediate filaments (IFs), together with actin filaments and microtubules, form the cytoskeleton &#8211; a critical structural element of every cell. ...

Measurement of Mitochondrial Oxygen Consumption in Permeabilized Fibers of Drosophila Using Minimal Amounts of Tissue

Medição do consumo de oxigênio mitocondrial nas fibras Permeabilized da drosófila usando quantidades mínimas de tecido

The fruit fly, Drosophila melanogaster, represents an emerging model for the study of metabolism. Indeed, drosophila have structures homologous to human ...

Study on the Metabolism of Six Systemic Insecticides in a Newly Established Cell Suspension Culture Derived from Tea (Camellia Sinensis L.) Leaves

Study on the Metabolism of Six Systemic Insecticides in a Newly Established Cell Suspension Culture Derived from Tea Camellia Sinensis L. Leaves

Estudo sobre o metabolismo de seis inseticidas sistêmicas em uma cultura de suspensão celular recém-estabelecida derivada do chá (Camellia sinensis L.) Folhas

A platform for studying insecticide metabolism using in vitro tissues of tea plant was developed. Leaves from sterile tea plantlets were induced to form ...

Isolation of Histone from Sorghum Leaf Tissue for Top Down Mass Spectrometry Profiling of Potential Epigenetic Markers

Isolamento de Histone do tecido da folha de sorgo para perfil de espectrometria de massa de top down de potenciais marcadores epigenéticos

Histones belong to a family of highly conserved proteins in eukaryotes. They pack DNA into nucleosomes as functional units of chromatin. ...

Preparation of Sample Support Films in Transmission Electron Microscopy using a Support Floatation Block

Preparação de filmes de suporte à amostra em microscopia eletrônica de transmissão usando um bloco de flutuação de suporte

Structure determination by cryo-electron microscopy (cryo-EM) has rapidly grown in the last decade; however, sample preparation remains a significant ...

Extraction and Purification of FAHD1 Protein from Swine Kidney and Mouse Liver

Extração e Purificação da Proteína FAHD1 do fígado de rim suíno e rato

Fumarylacetoacetate hydrolase domain-containing protein 1 (FAHD1) is the first identified member of the FAH superfamily in eukaryotes, acting as ...

Routine Collection of High-Resolution cryo-EM Datasets Using 200 KV Transmission Electron Microscope

Coleta rotineira de conjuntos de dados crio-EM de alta resolução usando microscópio eletrônico de transmissão de 200 KV

Cryo-electron microscopy (cryo-EM) has been established as a routine method for protein structure determination during the past decade, taking an ...

MyDoBID- A Novel Technique for Identifying Muscle Fiber Types Efficiently

Fiber Type Identification of Human Skeletal Muscle

Author Spotlight: Deciphering the Mysteries of Skeletal Muscle Fiber Types Using the MyDoBID Technique 

Identificação do Tipo de Fibra do Músculo Esquelético Humano

The technique described here can be used to identify specific myosin heavy chain (MHC) isoforms in segments of individual muscle fibers using dot ...

Accelerating Macromolecular Structural Analysis Using Smart Leginon Autoscreen

Cryo-Electron Microscopy Screening Automation Across Multiple Grids Using Smart Leginon

Author Spotlight: Enhancing Cryo-Electron Microscopy by Automated Data Collection and Analysis Techniques

Advancements in cryo-electron microscopy (cryoEM) techniques over the past decade have allowed structural biologists to routinely resolve macromolecular ...