Name: taste exam: a brief and validated test
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Awards from the National Institutes of Health supported this research (R01DC013588 to NAC, R21DC013886 to NAC and DRR, and NIDCD Administrative Research Supplement to Promote Emergence of Clinician-Scientists in Chemosensory Research to JED). We collected genotype data from equipment purchased in part with NIH funds from OD018125.

The University of Pennsylvania Institutional Review board approved this protocol. We excluded subjects if they were under 18 years of age or were pregnant.
1. Oral Cavity Evaluation: Disease Assessment and Papilla Identification
<ol>
	<li>Instruct the subject to open the mouth.</li>
	<li>Using a light source such as a penlight or headlamp, illuminate the oral cavity and examine the seven subsites of the area (tongue, floor of mouth, buccal mucosa, labial mucosa, gingiva, hard palate,...

<ol>
	<li>Cowart, B. J., Young, I. M., Feldman, R. S., Lowry, L. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clinical+disorders+of+smell+and+taste.">Clinical disorders of smell and taste.</a> Occupational Medicine. 12 (3), 465-483 (1997).</li><li>Ackerman, B. H., Kasbekar, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Disturbances+of+taste+and+smell+induced+by+drugs.">Disturbances of taste and smell induced by drugs.</a> Pharmacotherapy. 17 (3), 482-496 (1997).</li><li>Kveton, J. F., Bartoshuk, L. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+unilateral+chorda+tympani+damage+on+taste.">The effect of unilateral chorda tympani damage on taste.</a> Laryngoscope. 104, 25-29 (1994).</li><li>Fischer, R. A., Griffin, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pharmacogenetic+aspects+of+gustation.">Pharmacogenetic aspects of gustation.</a> Drug Research. 14 (14), 673-686 (1964).</li><li>Joyce, C. R., Pan, L., Varonos, D. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Taste+sensitivity+may+be+used+to+predict+pharmacological+effects.">Taste sensitivity may be used to predict pharmacological effects.</a> Life Science. 7 (9), 533-537 (1968).</li><li>Adappa, N. D., 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=Genetics+of+the+taste+receptor+T2R38+correlates+with+chronic+rhinosinusitis+necessitating+surgical+intervention.">Genetics of the taste receptor T2R38 correlates with chronic rhinosinusitis necessitating surgical intervention.</a> International Forum of Allergy &amp; Rhinology. , (2013).</li><li>Lipchock, S. V., Reed, D. R., Mennella, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Relationship+between+bitter-taste+receptor+genotype+and+solid+medication+formulation+usage+among+young+children:+a+retrospective+analysis.">Relationship between bitter-taste receptor genotype and solid medication formulation usage among young children: a retrospective analysis.</a> Clinical Therapeutics. 34 (3), 728-733 (2012).</li><li>Bartoshuk, L. M., Carterette, E. C., Friedman, M. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Handbook of perception: Tasting and smelling. , 2-18 (1978).</li><li>Coldwell, S. 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F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glucose+transporters+and+ATP-gated+K++(KATP)+metabolic+sensors+are+present+in+type+1+taste+receptor+3+(T1r3)-expressing+taste+cells.">Glucose transporters and ATP-gated K+ (KATP) metabolic sensors are present in type 1 taste receptor 3 (T1r3)-expressing taste cells.</a> Proceedings of the National Academy of Sciences USA. , (2011).</li><li>Tordoff, M. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Calcium:+taste,+intake+and+appetite.">Calcium: taste, intake and appetite.</a> Physiological Review. 81, 1567-1597 (2001).</li><li>Reed, D. R., Xia, M. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recent+advances+in+fatty+acid+perception+and+genetics.">Recent advances in fatty acid perception and genetics.</a> Advances in Nutrition. 6 (3), 353S-360S (2015).</li><li>Blekhman, 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=Host+genetic+variation+impacts+microbiome+composition+across+human+body+sites.">Host genetic variation impacts microbiome composition across human body sites.</a> Genome Biology. 16, 191 (2015).</li><li>Hoon, M. 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=Putative+mammalian+taste+receptors:+a+class+of+taste-specific+GPCRs+with+distinct+topographic+selectivity.">Putative mammalian taste receptors: a class of taste-specific GPCRs with distinct topographic selectivity.</a> Cell. 96 (4), 541-551 (1999).</li><li>Laffitte, A., Neiers, F., Briand, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+roles+of+the+sweet+taste+receptor+in+oral+and+extraoral+tissues.">Functional roles of the sweet taste receptor in oral and extraoral tissues.</a> Current Opinion in Clinical Nutrition and Metabolic. 17 (4), 379-385 (2014).</li><li>An, S. 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=Tas2r+activation+promotes+airway+smooth+muscle+relaxation+despite+beta2-adrenergic+receptor+tachyphylaxis.">Tas2r activation promotes airway smooth muscle relaxation despite beta2-adrenergic receptor tachyphylaxis.</a> American Journal of Physiology-Lung Cellular and Molecular. , (2012).</li><li>Fox, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+relationship+between+chemical+composition+and+taste.">The relationship between chemical composition and taste.</a> Science. 74, 607 (1931).</li><li>Fox, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+relationship+between+chemical+constitution+and+taste.">The relationship between chemical constitution and taste.</a> Proceedings of the National Academy of Sciences USA. 18, 115-120 (1932).</li><li>Bufe, B., 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=The+molecular+basis+of+individual+differences+in+phenylthiocarbamide+and+propylthiouracil+bitterness+perception.">The molecular basis of individual differences in phenylthiocarbamide and propylthiouracil bitterness perception.</a> Current Biol.ogy. 15 (4), 322-327 (2005).</li><li>Reed, D. 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=The+perception+of+quinine+taste+intensity+is+associated+with+common+genetic+variants+in+a+bitter+receptor+cluster+on+chromosome+12.">The perception of quinine taste intensity is associated with common genetic variants in a bitter receptor cluster on chromosome 12.</a> Human Molecular Genetics. 19 (21), 4278-4285 (2010).</li><li>Bobowski, N., Reed, D. R., Mennella, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Variation+in+the+TAS2R31+bitter+taste+receptor+gene+relates+to+liking+for+the+nonnutritive+sweetener+Acesulfame-K+among+children+and+adults.">Variation in the TAS2R31 bitter taste receptor gene relates to liking for the nonnutritive sweetener Acesulfame-K among children and adults.</a> Science Reports. 6, 39135 (2016).</li><li>Allen, A. L., McGeary, J. E., Knopik, V. S., Hayes, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bitterness+of+the+non-nutritive+sweetener+acesulfame+potassium+varies+with+polymorphisms+in+TAS2R9+and+TAS2R31.">Bitterness of the non-nutritive sweetener acesulfame potassium varies with polymorphisms in TAS2R9 and TAS2R31.</a> Chemical Senses. 38 (5), 379-389 (2013).</li><li>Roudnitzky, N., 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=Genomic,+genetic,+and+functional+dissection+of+bitter+taste+responses+to+artificial+sweeteners.">Genomic, genetic, and functional dissection of bitter taste responses to artificial sweeteners.</a> Human Molecular Genetics. 20 (17), 3437-3449 (2011).</li><li>Guo, S. W., Reed, D. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+genetics+of+phenylthiocarbamide+perception.">The genetics of phenylthiocarbamide perception.</a> Annals in Human Biology. 28 (2), 111-142 (2001).</li><li>Bartoshuk, L. 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=Labeled+scales+(e.g.,+category,+Likert,+VAS)+and+invalid+across-group+comparisons:+what+we+have+learned+from+genetic+variation+in+taste.">Labeled scales (e.g., category, Likert, VAS) and invalid across-group comparisons: what we have learned from genetic variation in taste.</a> Food Quality Prefererences. 14 (2), 125-138 (2003).</li></ol>

The significance of this method is that it uses a multidisciplinary approach with features from medicine (the oral exam), experimental psychology (the taste test) and genetics (a validation step). Taste information is likely to develop as a diagnostic and prognostic tool because taste provides a window into the function of proteins elsewhere in the body. From an experimental psychology viewpoint, the addition of a simple exam can identify subjects who are not appropriate for the study of normative taste function. From a ...

Measures of human taste perception can be both part of medical care and a target of biomedical research, yet taste has received scant attention compared with hearing and vision (Table 1). From the medical perspective, when clinicians evaluate patients complaining of taste loss, in most cases the actual loss is of smell1, which has led to dismissal of taste loss as an uncommon and often invalid presenting complaint. Taste distortions (dysgeusia) are more common and frequently arise from the secondary effects of medications or peripheral nerve injury2,3, but neither form has an effective treatment (other than stopping the medication). Clinicians have also ignored taste loss because it has hitherto had little diagnostic or prognostic value on its own. However, although the measurement of taste has been a backwater, it may now be entering mainstream medicine with the revival of a historical appreciation that taste may be a diagnostic or prognostic tool4,5. For instance, bitterness perception can predict immune function6 or the willingness of a patient to take medication7. Nonetheless, biomedical researchers have largely neglected taste. This inattention may, in part, reflect the fact that early progress in understanding this sensory system has its roots in experimental psychology8, a field with which those in medicine may be relatively unfamiliar. Moreover, renewed interest in taste has ushered in standardized taste methods9 that build on earlier methods10, which while comprehensive are lengthy and inappropriate for clinical settings. Finally, confidence in taste measures can be weak because subjects report on their own experience and validation of their observations has hitherto been lacking. Our hope is that a simple measure that investigators or clinicians can easily administer will gain in popularity with both constituents. Here we describe a simple taste exam protocol that has three parts: an assessment of the oral cavity, the taste test, and a validation step using inborn genotype. First, we provide biological context for these procedures, which merge simple practices in medicine, sensory measures from experimental psychology and validation of responses using genotype and genetics.
Taste perception starts in the mouth, so an effective taste exam needs to include a brief clinical assessment for obvious oral diseases, redness, swelling and other discoloration. The oral cavity contains seven subsites: the tongue, gingiva, floor of mouth, buccal mucosa, labial mucosa, hard palate, and the retromolar trigone. Past studies of human taste focused on healthy participants or those with well-defined diseases, but as taste testing becomes routine in medical exams, it is important to record the condition of the oral cavity as part of the procedure.
The tongue itself is a muscular structure encased in mucosa; dotting its dorsal surface are papillae, the small raised structures that give the tongue its unique texture and contain taste receptor cells. We classify papillae by their shape: fungiform, filiform, foliate, and circumvallate. Fungiform papillae (FP) are located anterolaterally on the tongue and are round, with a mushroom shape11. Investigators have published several useful methods to quantify FP and we direct readers to these sources for measurement protocols12,13,14,15,16. Foliate papillae, shaped like the pages of a book (folia), are located exclusively on the lateral posterior tongue surface11. Circumvallate papillae, found in the sulcus terminalis of the tongue base, are large dome-shaped structures surrounded by mucosal walls (Latin circum, &#34;surround,&#34; + vallum, &#34;wall&#34;)11. The most numerous papillae, the filiform, are long and thin and do not contain taste receptors.
People differ in tongue anatomy. While the sources of this anatomic variation are unknown, it is determined in part by inborn genetic variation, with investigators reporting 31% concordance of tongue anatomy among dizygotic twins and 60% concordance among monozygotic twins17. Papillary density also differs among people, and although rare, at least one genetic disease (familial dysautonomia) results in a congenital absence of taste papillae18,19,20. Thus, before performing psychophysical testing, it is helpful to confirm the presence of FP as part of the brief assessment and note the relative size and color of the tongue and evidence of oral disease.
The taste papillae contain the sensory cells that when stimulated initiate taste sensation. Humans are capable of sensing at least five classes of taste: salty, sour, bitter, sweet, and umami. While salty, sweet, and umami tastes signal the presence of valuable food sources containing sodium chloride, glucose, and amino acids, respectively, bitterness and sourness signal the presence of potential toxins and acids from the bacterial decomposition of food, respectively, and induce aversive behavior21. Salty and sour tastes are transduced through the activation of ion channels found in some types of taste cells, though the understanding of salt transduction is evolving and it may require type I cells as well22,23. Bitter, sweet, and umami arise from activation of G-protein-coupled receptors on type II taste cells, each attuned to a particular taste. Heterodimers of subunits of three particular receptors transduce sweet and umami while bitter compounds activate a group of 25 different bitter receptors24. These bitter receptors can respond to multiple bitter compounds, and a single bitter compound often stimulates more than one receptor25. Despite the recent expansion of knowledge about the molecular basis of taste, novel pathways26 and new discoveries beyond the traditional five taste qualities (e.g., calcium27 or fatty acid28 perception) may lie ahead.
There are at least two surprising aspects of the taste families of receptors: genes that code for these receptors can differ markedly in DNA sequence and hence function among people, and many tissues of the body express these genes21,29,30,31. These extraoral sites include the brain, thyroid, upper and lower respiratory tract, and the gastrointestinal tract, among many others21,29,30,31. While the taste receptors at these locations do not participate in taste perception in the traditional sense, they likely sense the local chemical environment29,32. For example, the ciliated epithelium of the upper respiratory tract expresses the bitter receptor T2R38 (Bitter Taste Receptor 38), which responds to chemical compounds produced by bacteria and influences the innate immune response32, such as increasing mucociliary clearance and levels of anti-microbial peptides and nitric oxide. This finding has medical implications for chronic rhinosinusitis, a disease of chronic bacterial infection and inflammation of the upper respiratory tract and paranasal sinuses.
Of particular relevance to the taste exam we describe here is that the T2R38 bitter taste receptor, encoded by the TAS2R38 gene, exhibits genetic variability and therefore variable taste sensitivity. Perceptual differences for the bitter compound phenylthiocarbamide (PTC) were first described by the chemist Arthur Fox33; later this compound was identified as an agonist of the T2R38 receptor34. Individual differences arise from the DNA sequence of the TAS2R38 gene, which has three single-nucleotide polymorphisms, each yielding amino acid substitutions (A49P, A262V, and I296V; A: Alanine, P: Proline, V: Valine, I: Isoleucine). Two common haplotypes result, PAV and AVI, with PAV/PAV individuals being highly sensitive to PTC (&#34;tasters&#34;), AVI/AVI individuals being relatively insensitive (&#34;non-tasters&#34;), and heterozygous AVI/PAV individuals being more variable in their sensitivity35. There are more examples of genetic variation affecting bitter perception, e.g., taste receptor T2R19, encoded by the TAS2R19 gene, similarly exhibits genetic variability and differing taste sensitivity to the bitter compound quinine36. Likewise, variation in TAS2R31 affects the perceived bitterness of one of the high-potency sweeteners37,38,39.
Here we describe a rapid method to characterize a patient&#39;s sense of taste that draws on high-yield protocols in clinical medicine, experimental psychology and genetics.

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			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:307px;">Phenylthiocarbamide</td>
			<td>Sigma Aldrich</td>
			<td>P7629</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:307px;">Quinine hydrochloride dihydrate</td>
			<td>Sigma Aldrich</td>
			<td>Q1125</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:307px;">Sodium Chloride</td>
			<td>Sigma Aldrich</td>
			<td>S1679</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:307px;">Sucrose</td>
			<td>Sigma Aldrich</td>
			<td>S0389</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Glass scintillation vials</td>
			<td>Thomas Scientific</td>
			<td>1230L59</td>
			<td>Same as Wheaton catalog no. 986580</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Oragene Discover OGR-500 DNA collection kit</td>
			<td>DNA Genotek</td>
			<td style="width:124px;">OGR-500</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">prepIT L2P Protocol reagents</td>
			<td>DNA Genotek</td>
			<td>PT-L2P-5</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">rs713598 TaqMan SNP genotyping assay</td>
			<td>ThermoFisher Scientific</td>
			<td>C___8876467_10</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">rs1726866 TaqMan SNP genotyping assay</td>
			<td>ThermoFisher Scientific</td>
			<td>C___9506827_10</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">rs10246939 TaqMan SNP genotyping assay</td>
			<td>ThermoFisher Scientific</td>
			<td>C___9506826_10</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">rs10772420 TaqMan SNP genotyping assay</td>
			<td>ThermoFisher Scientific</td>
			<td>C___1317426_10</td>
			<td></td>
		</tr>
	</tbody>
</table>

taste exam: a brief and validated test

The emerging importance of taste in medicine and biomedical research, and new knowledge about its genetic underpinnings, has motivated us to supplement classic taste-testing methods in two ways. First, we explain how to do a brief assessment of the mouth, including the tongue, to ensure that taste papillae are present and to note evidence of relevant disease. Second, we draw on genetics to validate taste test data by comparing reports of perceived bitterness intensity and inborn receptor genotypes. Discordance between objective measures of genotype and subjective reports of taste experience can identify data collection errors, distracted subjects or those who have not understood or followed instructions. Our expectation is that fast and valid taste tests may persuade researchers and clinicians to assess taste regularly, making taste testing as common as testing for hearing and vision. Finally, because many tissues of the body express taste receptors, taste responses may provide a proxy for tissue sensitivity elsewhere in the body and, thereby, serve as a rapid, point-of-care test to guide diagnosis and a research tool to evaluate taste receptor protein function.

Results from the taste test have been pooled for all subjects evaluated (n = 840) and are presented after segregation by genotype. The full data set is accessible at https://carayata.shinyapps.io/TasteBoxplots/ and can be reviewed for each tastant assessed and for TAS2R38 and TAS2R19 genotypes. Results confirm the existence of perceptual taste differences for PTC among subjects grouped by TAS2R38 receptor genotype (Figure 3). Rating...

Watch this Scientific Journal Video about Taste Exam: A Brief and Validated Test at JoVE.com

Taste Exam: A Brief and Validated Test

This protocol measures human taste responses and includes a brief anatomical assessment, a short taste test, and a validation method using the subject&#39;s reported sensation and taste receptor genotype.

The emerging importance of taste in medicine and biomedical research, and new knowledge about its genetic underpinnings, has motivated us to supplement ...

This method can help answer key questions in the medical and sensory field about each person's sense of taste. The main advantage of this technique is its brevity and suitability for biomedical research. The implications of this technique extend towards the diagnosis and treatment of chronic rhino sinusitis because taste receptors are part of the innate immune system.I will be demonstrating the procedure on our lab member David and Lauren will be making the taste solutions. Begin by instructing the subject to open their mouth. Use a light to illuminate the oral cavity and examine the seven sub-sites of the area.Here the dorsal tongue is easily visible. Using a tongue depressor to depress the tongue the posterior pharynx becomes visible. Next lateralizing the tongue allows exposure of the left retromolar trigone and the left buccal mucosa.Here the anterior gingiva and the labial mucosa are visualized. As the subject's tongue is lifted the ventral tongue surface becomes visible. And this allows for examination of the subjects floor of mouth.The tongue is then lateralized again to examine the right retromolar trigone and the right buccal mucosa. And lastly the hard and soft palate are visualized. Note lesions, abrasions, and masses or signs of inflammation.Use a light source to visualize the dorsal surface of the tongue and identify the presence or absence of fungiform papillae or smooth surface of the tongue. Finally write down the results of oral cavity examination before proceeding with taste testing. Begin by preparing the taste solutions using a volumetric flask for each to ensure precision of concentrations.Dissolve samples using ultra pure water. Wrap any light sensitive solutions with foil to reduce exposure to light. Then store the taste solutions at four degrees Celsius.To identify common errors in solution preparation fill one tasting cup with the old solution and one with the new solution. Taste each solution to verify they are identical in strength. Next, label vial caps with a circular sticker according to the order of presentation and a color code.In order to match the vial labels with the labels on the taste questionnaire. Package samples by placing them into their respective carriers. Next prepare the taste questionnaire using a category scale for rating taste intensity and a forced choice for identifying the taste quality of each tastant.Lastly, place circular labels of the appropriate color and number next to the appropriate sample in the taste questionnaire. Provide the subject with the sample box, a bottle of water, an empty cup, pen, and taste questionnaire containing entries for 12 samples. Explain the testing procedure and inform the subject that they will be asked to rate both the intensity and quality of each tastant.Let them know that they may not experience all qualities. Start the test by asking the subject to rinse their mouth twice with water and spit it out in the cup provided. Have them pour all of sample one into their mouth and hold it there for five seconds before spitting the solution into the cup.Tell them not to gargle or swallow the solution. Then instruct the subject to circle one of the 13 vertical lines corresponding with the sample's intensity. On a scale of 0 to 12.From no intensity at all to extremely intense. And choose a single quality to describe the taste. Next ask the subject to rinse their mouth with water twice before proceeding to the next sample.Observe the subject tasting and rating sample one of water. Should the rating deviate from no intensity at all and no flavor, reiterate the questionnaire instructions before allowing the test to proceed. After all samples have been tasted review the finished questionnaire for completeness.Finally, score intensity ratings on a scale of 0 to 12 from the vertical lines that the subject circled. And average the two intensity ratings for each tastant for later analysis. Results of the taste test confirm the existence of perceptual taste differences for the bitter compound, Phenylthiocarbamide or PTC among subjects grouped by the TAS2R38 receptor genotype.Ratings of PTC intensity are significantly different across three genotypes. The motivation behind this study was to try to assess or to develop a rapid point of service exam that we could do in the clinic that would assess the function of this receptor that we believe plays a role in the development of chronic sinusitis. The idea here is could we use this taste test to predict which of our patients are at high risk for developing sinusitis and more importantly, which patients are good surgical candidates for us to do our sinus surgery on, or which ones will do poorly with our sinus surgery.So we're trying to use this taste test to prognosticate patient's sinal-nasal disease. Once that we went through all of the taste testing we confirmed that by genotyping the patients. And as you can see in figure three for the most part the patient's genotype follows the pattern of their phenotype.Meaning the more bitter they tasted they tended to be dominant for this receptor. However, there are some outliers. Showing that there are functional homozygotes, PAV/PAVs that said that they didn't taste PTC and on the flip side of that there were some nonfunctional AVI/AVI individuals that tasted PTC with bitterness.So that means that whatever taste test we do develop if there are patients that are not fulfilling our expectations that we probably wanna confirm the taste test by genotyping them for T2R38. Once mastered the exam and taste test can be performed in under 30 minutes. After watching this video you should have an understanding of how to perform an oral exam, note evidence of disease and perform a brief taste test.

Research

JoVE Journal

Medicine

Making Sense of Listening: The IMAP Test Battery

The ability to hear is only the first step towards making sense of the range of information contained in an auditory signal. Of equal importance are the abilities to extract and use the information encoded in the auditory signal. We refer to these as listening skills (or auditory processing AP). Deficits in these skills are associated with delayed language and literacy development, though the nature of the relevant deficits and their causal connection with these delays is hotly debated.
When a child is referred to a health professional with normal hearing and unexplained difficulties in listening, or associated delays in language or literacy development, they should ideally be assessed with a combination of psychoacoustic (AP) tests, suitable for children and for use in a clinic, together with cognitive tests to measure attention, working memory, IQ, and language skills. Such a detailed examination needs to be relatively short and within the technical capability of any suitably qualified professional. Current tests for the presence of AP deficits tend to be poorly constructed and inadequately validated within the normal population. They have little or no reference to the presenting symptoms of the child, and typically include a linguistic component. Poor performance may thus reflect problems with language rather than with AP. To assist in the assessment of children with listening difficulties, pediatric audiologists need a single, standardized child-appropriate test battery based on the use of language-free stimuli.
We present the IMAP test battery which was developed at the MRC Institute of Hearing Research to supplement tests currently used to investigate cases of suspected AP deficits. IMAP assesses a range of relevant auditory and cognitive skills and takes about one hour to complete. It has been standardized in 1500 normally-hearing children from across the UK, aged 6-11 years. Since its development, it has been successfully used in a number of large scale studies both in the UK and the USA. IMAP provides measures for separating out sensory from cognitive contributions to hearing. It further limits confounds due to procedural effects by presenting tests in a child-friendly game-format. Stimulus-generation, management of test protocols and control of test presentation is mediated by the IHR-STAR software platform. This provides a standardized methodology for a range of applications and ensures replicable procedures across testers. IHR-STAR provides a flexible, user-programmable environment that currently has additional applications for hearing screening, mapping cochlear implant electrodes, and academic research or teaching.

A test battery (IMAP) for performing an in-depth assessment of auditory and cognitive abilities contributing to listening skills is described. It is quick to administer, child-friendly and free from linguistic confounds. Stimulus generation and protocol management are controlled via a software platform (IHR-STAR) to ensure replicable procedures.

The ability to hear is only the first step towards making sense of the range of information contained in an auditory signal. Of equal importance are the ...

Technique to Collect Fungiform (Taste) Papillae from Human Tongue

The sense of taste is critical for human life. It informs the body about the quality of food that will be potentially ingested and stimulates metabolic processes that prepare the alimentary canal for digestion. Steady progress is being made towards understanding the early biochemical and molecular events underlying taste transduction (for a review, Breslin and Spector, 20081). However, progress to date has largely resulted from animal models. Yet, since marked differences in receptor specificity and receptor density vary among species, human taste transduction will only be understood by using human taste tissue. Here we describe a biopsy technique to collect human fungiform papillae, visible as rounded pink anterior structures, about 0.5 mm in diameter that contain taste buds. These biopsied papillae are used for several purposes including the isolation of viable taste bud cells, in situ hybridization, immunohistochemistry and, through techniques of molecular biology, the identification of taste-specific novel proteins.

Technique to Collect Fungiform Taste Papillae from Human Tongue

Knowledge of molecular mechanisms underlying gustatory transduction has recently enjoyed significant advances, largely due to using animal models. However, the wide diversity in taste sensitivity and specificity among mammals warrants studies in human tissue. We describe a biopsy technique to collect living taste cells from the papillae on human tongue.

The sense of taste is critical for human life. It informs the body about the quality of food that will be potentially ingested and stimulates metabolic ...

The Trier Social Stress Test Protocol for Inducing Psychological Stress

This article demonstrates a psychological stress protocol for use in a laboratory setting. Protocols that allow researchers to study the biological pathways of the stress response in health and disease are fundamental to the progress of research in stress and anxiety.1 Although numerous protocols exist for inducing stress response in the laboratory, many neglect to provide a naturalistic context or to incorporate aspects of social and psychological stress. Of psychological stress protocols, meta-analysis suggests that the Trier Social Stress Test (TSST) is the most useful and appropriate standardized protocol for studies of stress hormone reactivity.2 In the original description of the TSST, researchers sought to design and evaluate a procedure capable of inducing a reliable stress response in the majority of healthy volunteers.3 These researchers found elevations in heart rate, blood pressure and several endocrine stress markers in response to the TSST (a psychological stressor) compared to a saline injection (a physical stressor).3 Although the TSST has been modified to meet the needs of various research groups, it generally consists of a waiting period upon arrival, anticipatory speech preparation, speech performance, and verbal arithmetic performance periods, followed by one or more recovery periods. The TSST requires participants to prepare and deliver a speech, and verbally respond to a challenging arithmetic problem in the presence of a socially evaluative audience.3 Social evaluation and uncontrollability have been identified as key components of stress induction by the TSST.4 In use for over a decade, the goal of the TSST is to systematically induce a stress response in order to measure differences in reactivity, anxiety and activation of the hypothalamic-pituitary-adrenal (HPA) or sympathetic-adrenal-medullary (SAM) axis during the task.1 Researchers generally assess changes in self-reported anxiety, physiological measures (e.g. heart rate), and/or neuroendocrine indices (e.g. the stress hormone cortisol) in response to the TSST. Many investigators have adopted salivary sampling for stress markers such as cortisol and alpha-amylase (a marker of autonomic nervous system activation) as an alternative to blood sampling to reduce the confounding stress of blood-collection techniques. In addition to changes experienced by an individual completing the TSST, researchers can compare changes between different treatment groups (e.g. clinical versus healthy control samples) or the effectiveness of stress-reducing interventions.1

This article describes a protocol for inducing psychological stress in participants, which enables researchers to measure psychological, physiological and neuroendocrine responses to stress within single participants or between groups.

This article demonstrates a psychological stress protocol for use in a laboratory setting.  Protocols that allow researchers to study the biological ...

The &alpha;-test: Rapid Cell-free CD4 Enumeration Using Whole Saliva

There is an urgent need for affordable CD4 enumeration to monitor HIV disease. CD4 enumeration is out of reach in resource-limited regions due to the time and temperature restrictions, technical sophistication, and cost of reagents, in particular monoclonal antibodies to measure CD4 on blood cells, the only currently acceptable method. A commonly used cost-saving and time-saving laboratory strategy is to calculate, rather than measure certain blood values. For example, LDL levels are calculated using the measured levels of total cholesterol, HDL, and triglycerides1. Thus, identification of cell-free correlates that directly regulate the number of CD4+ T cells could provide an accurate method for calculating CD4 counts due to the physiological relevance of the correlates.
The number of stem cells that enter blood and are destined to become circulating CD4+ T cells is determined by the chemokine CXCL12 and its receptor CXCR4 due to their influence on locomotion2. The process of stem cell locomotion into blood is additionally regulated by cell surface human leukocyte elastase (HLECS) and the HLECS-reactive active &alpha;1proteinase inhibitor (&alpha;1PI, &alpha;1antitrypsin, SerpinA1)3. In HIV-1 disease, &alpha;1PI is inactivated due to disease processes 4. In the early asymptomatic categories of HIV-1 disease, active &alpha;1PI was found to be below normal in 100% of untreated HIV-1 patients (median=12 &mu;M, and to achieve normal levels during the symptomatic categories4, 5. This pattern has been attributed to immune inactivation, not to insufficient synthesis, proteolytic inactivation, or oxygenation. We observed that in HIV-1 subjects with &gt;220 CD4 cells/&mu;l, CD4 counts were correlated with serum levels of active &alpha;1PI (r2=0.93, p&lt;0.0001, n=26) and inactive &alpha;1PI (r2=0.91, p&lt;0.0001, n=26) 5. Administration of &alpha;1PI to HIV-1 infected and uninfected subjects resulted in dramatic increases in CD4 counts suggesting &alpha;1PI participates in regulating the number of CD4+ T cells in blood 3.
With stimulation, whole saliva contains sufficient serous exudate (plasma containing proteinaceous material that passes through blood vessel walls into saliva) to allow measurement of active &alpha;1PI and the correlation of this measurement is evidence that it is an accurate method for calculating CD4 counts. Briefly, sialogogues such as chewing gum or citric acid stimulate the exudation of serum into whole mouth saliva. After stimulating serum exudation, the activity of serum &alpha;1PI in saliva is measured by its capacity to inhibit elastase activity. Porcine pancreatic elastase (PPE) is a readily available inexpensive source of elastase. PPE binds to &alpha;1PI forming a one-to-one complex that prevents PPE from cleaving its specific substrates, one of which is the colorimetric peptide, succinyl-L-Ala-L-Ala-L-Ala-p-nitroanilide (SA3NA). Incubating saliva with a saturating concentration of PPE for 10 min at room temperature allows the binding of PPE to all the active &alpha;1PI in saliva. The resulting inhibition of PPE by active &alpha;1PI can be measured by adding the PPE substrate SA3NA. (Figure 1). Although CD4 counts are measured in terms of blood volume (CD4 cells/&mu;l), the concentration of &alpha;1PI in saliva is related to the concentration of serum in saliva, not to volume of saliva since volume can vary considerably during the day and person to person6. However, virtually all the protein in saliva is due to serum content, and the protein content of saliva is measurable7. Thus, active &alpha;1PI in saliva is calculated as a ratio to saliva protein content and is termed the &alpha;1PI Index. Results presented herein demonstrate that the &alpha;1PI Index provides an accurate and precise physiologic method for calculating CD4 counts.

The &amp;#945;-test: Rapid Cell-free CD4 Enumeration Using Whole Saliva

A CD4 enumeration method, the &#x03B1;-test, is described which uses whole saliva to provide rapid and accurate CD4 counts. The &alpha;-test costs pennies and eliminates the need for technical training, costly reagents such as monoclonal antibodies, instrumentation, refrigeration, transport of samples, as well as collection and handling of blood.

There is an urgent need for affordable CD4 enumeration to monitor HIV disease. CD4 enumeration is out of reach in resource-limited regions due to the time ...

Flexible Colonoscopy in Mice to Evaluate the Severity of Colitis and Colorectal Tumors Using a Validated Endoscopic Scoring System

The use of modern endoscopy for research purposes has greatly facilitated our understanding of gastrointestinal pathologies. In particular, experimental endoscopy has been highly useful for studies that require repeated assessments in a single laboratory animal, such as those evaluating mechanisms of chronic inflammatory bowel disease and the progression of colorectal cancer. However, the methods used across studies are highly variable. At least three endoscopic scoring systems have been published for murine colitis and published protocols for the assessment of colorectal tumors fail to address the presence of concomitant colonic inflammation. This study develops and validates a reproducible endoscopic scoring system that integrates evaluation of both inflammation and tumors simultaneously. This novel scoring system has three major components: 1) assessment of the extent and severity of colorectal inflammation (based on perianal findings, transparency of the wall, mucosal bleeding, and focal lesions), 2) quantitative recording of tumor lesions (grid map and bar graph), and 3) numerical sorting of clinical cases by their pathological and research relevance based on decimal units with assigned categories of observed lesions and endoscopic complications (decimal identifiers). The video and manuscript presented herein were prepared, following IACUC-approved protocols, to allow investigators to score their own experimental mice using a well-validated and highly reproducible endoscopic methodology, with the system option to differentiate distal from proximal endoscopic colitis (D-PECS).

Over the past few years, new generation endoscopes have emerged as important diagnostic research aids for evaluating murine colitis and colorectal tumors. We present herein a detailed protocol for endoscopic assessment of inflammation and colorectal tumors in mice, as well as a novel scoring system that uses decimal identifiers to document the endoscopic severity of colitis and colorectal tumors.

The use of modern endoscopy for research purposes has greatly facilitated our understanding of gastrointestinal pathologies. In particular, experimental ...

The Multiple Sclerosis Performance Test (MSPT): An iPad-Based Disability Assessment Tool

Precise measurement of neurological and neuropsychological impairment and disability in multiple sclerosis is challenging. We report a new test, the Multiple Sclerosis Performance Test (MSPT), which represents a new approach to quantifying MS related disability. The MSPT takes advantage of advances in computer technology, information technology, biomechanics, and clinical measurement science. The resulting MSPT represents a computer-based platform for precise, valid measurement of MS severity. Based on, but extending the Multiple Sclerosis Functional Composite (MSFC), the MSPT provides precise, quantitative data on walking speed, balance, manual dexterity, visual function, and cognitive processing speed. The MSPT was tested by 51 MS patients and 49 healthy controls (HC). MSPT scores were highly reproducible, correlated strongly with technician-administered test scores, discriminated MS from HC and severe from mild MS, and correlated with patient reported outcomes. Measures of reliability, sensitivity, and clinical meaning for MSPT scores were favorable compared with technician-based testing. The MSPT is a potentially transformative approach for collecting MS disability outcome data for patient care and research. Because the testing is computer-based, test performance can be analyzed in traditional or novel ways and data can be directly entered into research or clinical databases. The MSPT could be widely disseminated to clinicians in practice settings who are not connected to clinical trial performance sites or who are practicing in rural settings, drastically improving access to clinical trials for clinicians and patients. The MSPT could be adapted to out of clinic settings, like the patient&#8217;s home, thereby providing more meaningful real world data. The MSPT represents a new paradigm for neuroperformance testing. This method could have the same transformative effect on clinical care and research in MS as standardized computer-adapted testing has had in the education field, with clear potential to accelerate progress in clinical care and research.

The Multiple Sclerosis Performance Test MSPT: An iPad-Based Disability Assessment Tool

Precise measurement of neurological and neuropsychological impairment and disability in multiple sclerosis is challenging. We report methodologic details on a new test, the Multiple Sclerosis Performance Test (MSPT). This new approach to the objective of quantification of MS related disability provides a computer-based platform for precise, valid measurement of MS severity.

Precise measurement of neurological and neuropsychological impairment and disability in multiple sclerosis is challenging. We report a new test, the ...

Determining The Electromyographic Fatigue Threshold Following a Single Visit Exercise Test

Theoretically, the electromyographic (EMG) fatigue threshold is the exercise intensity an individual can maintain indefinitely without the need to recruit more motor units which is associated with an increase in the EMG amplitude. Although different protocols have been used to estimate the EMG fatigue threshold they require multiple visits which are impractical for a clinical setting. Here, we present a protocol for estimating the EMG fatigue threshold for cycle ergometry which requires a single visit. This protocol is simple, convenient, and completed within 15-20 min, therefore, has the potential to be translated into a tool that clinicians can use in exercise prescription.

This protocol describes the electromyographic fatigue threshold which demarcates between nonfatiguing and fatiguing exercise workloads. This information could be used to develop a more individualized training program.

Theoretically, the electromyographic (EMG) fatigue threshold is the exercise intensity an individual can maintain indefinitely without the need to recruit ...

A Rat Carotid Balloon Injury Model to Test Anti-vascular Remodeling Therapeutics

The rat carotid balloon injury is a well-established surgical model that has been used to study arterial remodeling and vascular cell proliferation. It is also a valuable model system to test, and to evaluate therapeutics and drugs that negate maladaptive remodeling in the vessel. The injury, or barotrauma, in the vessel lumen caused by an inflated balloon via an inserted catheter induces subsequent neointimal growth, often leading to hyperplasia or thickening of the vessel wall that narrows, or obstructs the lumen. The method described here is sufficiently sensitive, and the results can be obtained in relatively short time (2 weeks after the surgery). The efficacy of the drug or therapeutic against the induced-remodeling can be evaluated either by the post-mortem pathological and histomorphological analysis, or by ultrasound sonography in live animals. In addition, this model system has also been used to determine the therapeutic window or the time course of the administered drug. These studies can leadto the development of a better administrative strategy and a better therapeutic outcome. The procedure described here provides a tool for translational studies that bring drug and therapeutic candidates from bench research to clinical applications.

The rat carotid balloon injury model described below allows researchers to evaluate drugs or therapeutics that negate injury-induced arterial hyperplasia. Detailed pre-surgical preparation, surgical procedure, and post-surgical cares of the animal are described.

The rat carotid balloon injury is a well-established surgical model that has been used to study arterial remodeling and vascular cell proliferation. It is ...

Ultrasound Assessment of Endothelial Function: A Technical Guideline of the Flow-mediated Dilation Test

Cardiovascular disease is the primary cause of mortality and a major cause of disability worldwide. The dysfunction of the vascular endothelium is a pathological condition characterized mainly by a disruption in the balance between vasodilator and vasoconstrictor substances and is proposed to play an important role in the development of atherosclerotic cardiovascular disease. Therefore, a precise evaluation of endothelial function in humans represents an important tool that could help better understand the etiology of multiple cardio-centric pathologies.
Over the past twenty-five years, many methodological approaches have been developed to provide an assessment of endothelial function in humans. Introduced in 1989, the FMD test incorporates a forearm occlusion and subsequent reactive hyperemia that promotes nitric oxide production and vasodilation of the brachial artery. The FMD test is now the most widely utilized, non-invasive, ultrasonic assessment of endothelial function in humans and has been associated with future cardiovascular events.
Although the FMD test could have clinical utility, it is a physiological assessment that has inherited several confounding factors that need to be considered. This article describes a standardized protocol for determining FMD including the recommended methodology to help minimize the physiological and technical issues and improve the precision and reproducibility of the assessment.

The flow mediated dilation (FMD) test is the most commonly utilized, non-invasive, ultrasound assessment of endothelial function in humans. Although the FMD test has been related with the prediction of future cardiovascular disease and events, it is a physiological assessment with many inherent confounding factors that need to be considered.

Cardiovascular disease is the primary cause of mortality and a major cause of disability worldwide. The dysfunction of the vascular endothelium is a ...

A Blood-based Test for the Detection of ROS1 and RET Fusion Transcripts from Circulating Ribonucleic Acid Using Digital Polymerase Chain Reaction

We have developed novel methods for the isolation and characterization of tumor-derived circulating ribonucleic acid (cRNA) for blood-based liquid biopsy. Robust detection of cRNA recovered from blood represents a solution to a critical unmet need in clinical diagnostics. The test begins with the collection of whole blood into blood collection tubes containing preservatives that stabilize cRNA. Cell-free, exosomal, and platelet-associated RNA is isolated from plasma in this test system. The cRNA is reverse transcribed to complementary DNA (cDNA) and amplified using digital polymerase chain reaction (dPCR). Samples are evaluated for both the target biomarker as well as a control gene. Test validation included limit of detection, accuracy, and robustness studies with analytic samples. The method developed as a result of these studies reproducibly detect multiple fusion variants for ROS1 (C-Ros proto-oncogene 1; 8 variants) and RET (rearranged during transfection proto-oncogene; 8 variants). The sample processing workflow has been optimized so that test results can consistently be generated within 72 hours of sample receipt.

Detection of circulating ribonucleic acid (cRNA) from blood is an unmet need in clinical diagnostics. Here we describe methods that characterize cRNA from non-small cell lung cancer patients using sensitive and specific digital polymerase chain reaction. The tests meet design requirements to detect fusion variants within 72 hours.

We have developed novel methods for the isolation and characterization of tumor-derived circulating ribonucleic acid (cRNA) for blood-based liquid biopsy. ...