The research and development leading to the IMAP-test battery and IHR-STAR was funded by the Medical Research Council (MRC) and a grant from the Oticon Foundation. Thanks go to Lorna Halliday, Sally Hind and Alison Riley, who provided help in the planning stages of the work and Sonja Ratib and team for testing the IMAP test battery during the large population-based study or auditory processing. IHR technical and support staff provided invaluable assistance with the project - we particularly acknowledge the contributions of Tim Folkard, Victor Chilekwa, Dave Bullock and John Chambers.

This protocol is aimed at both researchers and clinicians. It briefly outlines: (i) the IMAP ('IHR multicentre study of auditory processing') test battery which is based primarily on the use of nonverbal AP tests, (ii) information on the factors introduced into the battery to ensure a successful child-centric approach, and (iii) a step-by-step guide to describe the test procedure to be used with children aged six years or older.
1. IMAP test battery
The IMAP battery provides a comprehensive auditory and cognitive assessment which can be administered in about one hour. It comprises a total of 10 tests: five AP tests, based on non-linguistic materials, a speech-in-noise test, a sustained attention test, designed to assess attention in both the auditory and visual modalities, and four cognitive tasks that assess reading, verbal short-term memory, working memory and nonverbal intelligence (NVIQ). Brief descriptions of these tests are provided in Appendix A.
During testing, auditory stimuli are delivered through comfortable, high quality headphones (e.g. Sennheiser HD25) and the child responds via a colorful, three button box (Figure 1). The IMAP test battery is presented via the IHR-STAR (IHR System for Testing Auditory Responses) platform test. This software platform comprises a suite of programs for generating stimuli, controlling the order of presentation of AP and cognitive tests, and ensuring that test protocols are adhered to. 
 <img src="/files/ftp_upload/2139/2139fig1.jpg" alt="Figure 1" /> 
 Figure 1. A child being tested on the IMAP test-battery. The key features to notice are the colorful button box, the use of headphones through which sounds are presented, and the child-friendly images on the computer-screen. 
 

Two 'tracks' are presented for each AP test. The tracks each comprise 20 trials, where a trial is a sequence of three stimuli (inter-stimulus interval 400 ms): two identical 'standard' stimuli and a different, 'target' stimulus. The location of the target stimulus is varied randomly within the sequence and child is required to identify the 'odd-one-out'. This is called a three-interval, three-alternate forced-choice (3I-3AFC) oddball response paradigm (Figure 2). 
 
 <img src="/files/ftp_upload/2139/2139fig2.jpg" alt="Figure 2" /> 
 Figure 2. Illustration of a trial in a test of frequency discrimination. There are three intervals i.e., three tones. Two tones are the same (1000 Hz) while the third one is different (1500 Hz). This is the target. 
 

The standard tone in IMAP is a 1000 Hz for all tests. The parameters for the 'target' tone depend on the AP-test and details are provided in Table 1. During testing the relevant parameters for the 'target' are varied according to the child's previous response using an 'adaptive', staircase method which is designed to converge quickly on an estimate of threshold (i.e., minimum detectable difference). See Table 1 for a summary of the different rules for step-size changes for the AP-tests.
Stimulus and response data are stored electronically for later extraction and analysis.
<table border="1">
 <tbody>
 <tr>
 <td>Test</td>
 <td>Track rules (step size, staircase)</td>
 <td>Target frequency; duration; initial presentation level</td>
 <td>Masking noise type; centre frequency; width; level; duration</td>
 </tr>
 <tr>
 <td>Backward masking 
 (0 ms gap)</td>
 <td>1. 15 dB, 1-down 1-up 
 2. 10 dB, 1-down 1-up 
 3. 5 dB, 3-down 1-up 
 Linear scale</td>
 <td>1000 Hz; 20 ms (10 ms cosine-ramp); 
 
 90 dB SPL</td>
 <td>Bandpass; 1000 Hz; 800 Hz; 30 dB/Hz; 300 ms</td>
 </tr>
 <tr>
 <td>Backward masking 
 (50 ms gap)</td>
 <td>As above</td>
 <td>1000 Hz; 20 ms (10 ms cosine- ramp) 
 75 dB SPL</td>
 <td>As above</td>
 </tr>
 <tr>
 <td>Simultaneous masking (no-notch)</td>
 <td>As above</td>
 <td>1000 Hz; 20 ms (10 ms cosine-ramp) 
 95 dB SPL</td>
 <td>As above</td>
 </tr>
 <tr>
 <td>Simultaneous masking 
 ( notch)</td>
 <td>As above</td>
 <td>1000 Hz; 20 ms (10 ms cosine-ramp) 
 90 dB SPL</td>
 <td>Bandstop; 1000 Hz; 1200 Hz (400 Hz spectral notch); 30 dB/Hz ; 300 ms</td>
 </tr>
 <tr>
 <td>Frequency discrimination</td>
 <td>1. 2 &#948;Hz, 1-down 1-up 
 2. 2 &#948;Hz 1-down 1-up 
 3. 1.41&#948;Hz 3-down 1-up 
 Log scale</td>
 <td>S = 1000 Hz 
 T = 1500 Hz (&#948; = 50%) 
 200 ms; 70 dB SPL</td>
 <td>Not applicable</td>
 </tr>
 <tr>
 <td>VCV speech-in-noise</td>
 <td>1. 10 dB, 1-down 1-up 
 2. 5 dB, 1-down 1-up 
 3. 3 dB, 3-down 1-up</td>
 <td>T = consonant in VCV e.g., aGa 
 80 dBA</td>
 <td>ICRA-5 (male; one-speaker); 60 dBA</td>
 </tr>
 </tbody>
</table>
Table 1. Stimulus parameters and adaptive rules for the AP and VCV tests. S = standard; T = target.
2. Child-friendly Approach
Specific elements internal and external to the software are used to enhance and maximize the child's engagement with the protocol.
Communication. It is essential to establish a good rapport between the tester and the child. This includes using age-appropriate language, presenting the test session as a series of 'games to play' rather than 'tests to perform', talking directly to the child (not just to the parent) and using appropriate child-friendly body language. A standardized dialogue has been developed for each test (see Appendix B), which ensures instructions are clear and age appropriate. The tester must also check that the child has fully understood the instructions.
Test environment. Testing should be conducted in a quiet room with no auditory or visual distractions. This minimizes masking effects of background noise, and avoids confounds due to the child's attention being drawn away from the tests. Ideally, only the child and tester should be present during testing.
Familiarization. A familiarization phase is incorporated into the protocol. It is performed prior to testing, and includes presentation of easy-to-identify suprathreshold stimuli alongside impossible-to-detect stimuli. This ensures the oddball paradigm and forced-choice test formats are understood. The tester can then provide further clarification if necessary. Familiarization is further facilitated by beginning each test with easily identifiable target stimuli so the child enjoys an immediate sense of success.
Lapses of attention. An incorrect response early in the track due to a temporary lapse in attention can result in an artificially elevated threshold. To minimize this, the track is automatically abandoned when an incorrect response occurs within the first two trials. Tracks can be restarted a maximum of three times before testing is aborted for that particular test.
Minimization of boredom. AP track durations are designed to be short (20 trials) and each take between 1.5 and 2 minutes to complete. Listening and cognitive tests are interleaved within the test sequence to maintain a sense of novelty for the child. Finally, the 'forced-choice' method allows the child to work at his/her own pace.
Engagement. The tests are presented as fun, computer games to play. They include child-friendly graphics that change for each game. The cartoon animal or human characters move dynamically with each stimulus presentation and response (Figure 3). The child's attention is directed to each character and the sound that it makes by means of a 'bouncing ball'. The ball also indicates to the child when to respond. Auditory and visual feedback is given after each response, and there is an indicator to show how far the child has progressed through the 'game'. 
 
 <img src="/files/ftp_upload/2139/2139fig3.jpg" alt="Figure 3" /> 
 Figure 3. A screen shot from the IMAP test battery showing different features in the program interface that have been designed to maintain child engagement. 
 

Motivation and rewards. Although the procedural aspects of test presentation in the IMAP battery are controlled by IHR-STAR, it is essential for the tester to also engage with the child throughout testing. This involves providing positive motivational feedback between tests to engender a feeling of success. Cartoon stickers and certificates are given on completion of testing, and stickers may be used as rewards during testing if necessary.
Session Protocol
<ol>
 <li>For research studies, age-appropriate information sheets for both child and parent are sent out prior to the test session.</li>
 <li>The tester welcomes the child and the parent (if present) and briefly describes what will happen. For testing in a research context, informed written parental consent is obtained if it has not been obtained previously. The child's assent is also obtained.</li>
 <li>An anonymized participant ID number is entered into the appropriate box on the ID screen and this is then automatically appended to all data files collected as part of the test session.</li>
 <li>One out of 10 possible pseudo-randomized test sequences is automatically selected and testing begins.</li>
 <li>The tester introduces the first AP game display to the child and explains the test procedure prior to placing the headphones on the child and starting the familiarization phase for the test. This phase is completed automatically when a preset criterion (e.g. positive response to 4/5 trials) is reached. The child then enters the main testing phase.</li>
 <li>Tests are presented automatically according to the computer-controlled sequence. Prior to each test, the tester instructs the child as appropriate, and gives positive feedback and further encouragement as and when required.</li>
 <li>On completion of the test sequence, the child is rewarded with stickers and a certificate. If present, the parent is provided with a verbal summary of their child's results. These are compared against age-specific norms to account for developmental differences between children. Further management is discussed as appropriate.</li>
</ol>
3. Representative Results
Figure 4 provides examples of good and poor performances for two different children. In the good performance (left panel), the child consistently identifies the 'odd-one-out' and both tracks rapidly converge to the same threshold for intensity level. In other words, both the between- and within-track responses are highly consistent, with minimal deviation. This performance suggests on-task attention has been maintained throughout both tests.
The right panel shows response tracks from a younger child. It is clear, from the rapid convergence to threshold on the first track, that the child has understood the task. However variations in on-task attention are also apparent in both the first (blue) and second (red) tracks. In the first track, the child responds with reasonable consistency up to the last four trials where the child provides a succession of errors, resulting in a higher final threshold than might have been predicted earlier in the track. During the second track, the child fails to achieve performance comparable to the lowest level of detection that was observed in the earlier portion of the first run. There is a succession of six incorrect responses, leading to a progressive increase in the presentation level of the target up to approximately 40 dB above that of the first track. Correct responses then follow, at around trial 13, and the child continues with a series of correct responses until the track finishes. The large inter-track response variation, in conjunction with the large intra-track difference for the second track, suggests a lapse in attention rather than a problem with AP per se. 
 
 <img src="/files/ftp_upload/2139/2139fig4.jpg" alt="Figure 4" /> 
 Figure 4. Two representative graphs demonstrating of performance that can be obtained even when procedural effects are minimized. The left panel is an example of consistent reliable performance for both the first (blue) and second (red) tracks. The right panel provides evidence for lapses of attention after initially good performance (blue) track compared to the second (red) track.
<a href="https://www.jove.com/files/ftp_upload/2139/2139Appendix_A.pdf">Click here for Appendix A.</a> 
<a href="https://www.jove.com/files/ftp_upload/2139/2139Appendix_B.pdf">Click here for Appendix B.</a>

<ol>
	<li>Tallal, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fine-grained+discrimination+deficits+in+language-learning+impaired+children+are+specific+neither+to+the+auditory+modality+nor+to+speech+perception.">Fine-grained discrimination deficits in language-learning impaired children are specific neither to the auditory modality nor to speech perception.</a> J Speech, Lang, Hearing Res. 33, 616-621 (1990).</li><li>Tallal, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Improving+language+and+literacy+is+a+matter+of+time.">Improving language and literacy is a matter of time.</a> Nature Reviews: Neuroscience. 5, 721-728 (2004).</li><li>Wright, B. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Deficits+in+auditory+temporal+and+spectral+resolution+in+language-impaired+children.">Deficits in auditory temporal and spectral resolution in language-impaired children.</a> Nature. 387, 176-178 (2008).</li><li>Bishop, D. V. M., Carlyon, R. P., Deeks, J. M., Bishop, S. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Auditory+temporal+processing+impairment:+Neither+necessary+nor+sufficient+for+causing+language+impairment+in+children.">Auditory temporal processing impairment: Neither necessary nor sufficient for causing language impairment in children.</a> J Speech, Lang, Hearing Res. 42, 1295-1310 (1999).</li><li>Moore, D. R., Ferguson, M. A., Edmondson-Jones, A. M., Ratib, S., Riley, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The nature of auditory processing disorder in children. , (2010).</li><li>Strait, D. L., Kraus, N., Parbery-Clark, A., Ashley, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Musical+experience+shapes+top-down+auditory+mechanisms:+Evidence+from+masking+and+auditory+attention+performance.">Musical experience shapes top-down auditory mechanisms: Evidence from masking and auditory attention performance.</a> Hear Res. , (2009).</li><li>McArthur, G. M. &. a. m. p. ;. a. m. p., Bishop, D. V. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Frequency+discrimination+deficits+in+people+with+specific+language+impairment:+reliability,+validity,+and+linguistic+correlates.">Frequency discrimination deficits in people with specific language impairment: reliability, validity, and linguistic correlates.</a> J Speech, Lang, Hearing Res. 47, 527-541 (2004).</li><li>Ferguson, M. A., Hall, R. L., Riley, A., Moore, 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=Communication,+listening,+speech+and+cognition+in+children+with+auditory+processing+disorder+(APD)+or+specific+language+impairment+(SLI).">Communication, listening, speech and cognition in children with auditory processing disorder (APD) or specific language impairment (SLI).</a> J Speech, Lang, Hearing Res. , .</li><li>Wechsler, 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> WISC-IV(UK). , (2004).</li><li>Korkman, M., Kirk, U., Kemp, S. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> NEPSY: A developmental neuropsychological assessment. , (1998).</li><li>Bishop, D. V. M., North, T., Donlan, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nonword+repetition+as+a+behaviour+marker+for+inherited+language+impairment:+Evidence+from+a+twin+study.">Nonword repetition as a behaviour marker for inherited language impairment: Evidence from a twin study.</a> J Child Psychol Psyc. 37, 391-403 (1996).</li><li>Barry, J. G., Yasin, I., Bishop, D. V. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heritable+risk+factors+associated+with+language+impairments.">Heritable risk factors associated with language impairments.</a> Genes Brain Behav. 6, 66-76 (2007).</li><li>Wechsler, D., Chen, H. -. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Wechsler Abbreviated Scale of Intelligence. , (1999).</li><li>Torgesen, J. K., Wagner, R., Rashotte, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Test of Word Reading Efficiency (TOWRE). , (1999).</li><li>Bishop, D. V. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The Children's Communication Checklist-2. , (2003).</li><li>Smoski, W. J., Brunt, M. A., Tannahill, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Children's Auditory Processing Performance Scale. , (1998).</li></ol>

No conflicts of interest declared.

Good listening skills (i.e., AP) are believed to provide a firm basis for the further development of important cognitive skills such as speech, language and literacy1-3. Yet, there has been much debate about the nature of the relevant listening skills, if indeed they are relevant, necessary, or sufficient for causing difficulties in language or literacy4. Part of the problem for understanding the role of listening skills in cognitive development reflects the fact that children are very variable in their ability to perform taxing and, from their perspective, rather dull tests of AP ('psychophysical tasks'). This makes it difficult to separate out procedural effects, such as boredom, from more fundamental variations in functioning of central auditory processing pathways. As we have detailed in our protocol, the IMAP test battery was carefully designed to minimize confounds arising from issues associated with the test procedure. In Figure 4, we provide comparisons of performance of two children on a backward masking task. Because IMAP minimizes procedural confounds, and children cannot begin the test until they demonstrate they understand it, we can be confident in ascribing the different performances of these two children to test-specific factors for each child. The inclusion in the IMAP battery of paired AP tests5 and a range of cognitive tests further permits us to explore the reasons for this individual variation. We recently completed a UK population-based study using IMAP5. This showed how response variability and performance on tests tapping cognitive skills, such as attention, explained most of the variation in performance on the AP tests. The population-based study also provided us with standardized measures of performance. These measures are necessary to assess individual performance in the clinic. They are currently being used in a study investigating heritability of listening skills in identical and fraternal twins. This latter study will permit us to develop standardized measures suitable for use with children in the US. Though currently child-focused in design, IMAP can also be used with adults, as demonstrated by Strait et al.,6 who found an important contribution from cognitive skills honed by musical training to the development of auditory processing skills.

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		<div>
		<table border="1">
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		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>IHR-STAR2</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>presentation platform</td>
</tr>
<tr>
<td>IHR-Runner program</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>for pseudo-random order of presentation of tests</td>
</tr>
<tr>
<td>Laptop</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>Processor speed &#x2013; 2 GigaHz; 
RAM &#x2013; 1 Gigabyte; 
Operating system - Windows XP</td>
</tr>
<tr>
<td>USB IHR-audio-device</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>to control sound presentation level and spectrum content</td>
</tr>
<tr>
<td>Response button box</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Headphones</td>
<td>&nbsp;</td>
<td>Sennheiser</td>
<td>HD25-1</td>
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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.

Watch this Scientific Journal Video about Making Sense of Listening: The IMAP Test Battery at JoVE.com

Making Sense of Listening: The IMAP Test Battery

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 ...

making-sense-of-listening-the-imap-test-battery

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15.6K Views.  MRC Institute of Hearing Research. 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.

Video: Making Sense of Listening: The IMAP Test Battery

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 ...

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 ...

Characterization of Metabolic Status in Nonhuman Primates with the Intravenous Glucose Tolerance Test

The intravenous glucose tolerance test (IVGTT) plays a key role in the characterization of glucose homeostasis. When taken together with serum biochemical profiles, inclusive of blood glucose levels in both the fed and fasted state, HbA1c, insulin levels, clinical history of diet, body composition, and body weight status, an assessment of normal and abnormal glycemic control can be made. Interpretation of an IVGTT is done through measurement of changes in glucose and insulin levels over time in relation to the dextrose challenge. Critical components to be considered are: peak glucose and insulin levels reached in relation to T0 (end of glucose infusion), the glucose clearance rate K derived from the slope of rapid glucose clearance in the first 20 min (T1 to T20), the time to return to glucose baseline, and the area under the curve (AUC). These IVGTT measures will show characteristic changes as glucose homeostasis moves from a healthy to a diseased metabolic state5. Herein we will describe the characterization of nonhuman primates (Rhesus and Cynomolgus macaques), which are the most relevant animal model of Type II diabetes (T2D) in humans and the IVGTT and clinical profiles of these animals from a lean healthy, to obese dysmetabolic, and T2D state 8, 10, 11.

The goal of this protocol is to present a standard method to perform intravenous glucose tolerance tests (IVGTTs) to assess glycemic control in nonhuman primates and assess their metabolic status from healthy to dysmetabolic.

The intravenous glucose tolerance test (IVGTT) plays a key role in the characterization of glucose homeostasis. When taken together with serum biochemical ...

Design, Fabrication, and Administration of the Hand Active Sensation Test (HASTe)

The concept of personalizing neurologic rehabilitation, based on individual impairments, has experienced a recent surge. In parallel, the number of outcome measures of upper extremity motor performance has grown. However, clinicians and researchers lack practical, quantitative measures of the hand&#8217;s natural role as a receptor of the environment. The Hand Active Sensation Test (HASTe), developed by Williams and colleagues in 2006, is a valid and reliable measure of haptic performance. Though not available commercially, the HASTe can be fabricated from inexpensive materials, and it has been used successfully to identify impairments in haptic touch in individuals with stroke. (Williams, 2006). This paper presents the methods of design and fabrication of the HASTe testing kit, as well as a visual screen to be used during administration, and instructions for the tests administration and scoring.

Design, Fabrication, and Administration of the Hand Active Sensation Test HASTe

The Hand Active Sensation Test (HASTe) is a valid and reliable measure of haptic performance, which has been used successfully to identify impaired haptic touch in individuals with stroke. The purpose of this paper is to describe the design, fabrication and administration of the HASTe.

The concept of personalizing neurologic rehabilitation, based on individual impairments, has experienced a recent surge. In parallel, the number of ...

A Battery of Motor Tests in a Neonatal Mouse Model of Cerebral Palsy

As the sheer number of transgenic mice strains grow and rodent models of pediatric disease increase, there is an expanding need for a comprehensive, standardized battery of neonatal mouse motor tests. These tests can validate injury or disease models, determine treatment efficacy and/or assess motor behaviors in new transgenic strains. This paper presents a series of neonatal motor tests to evaluate general motor function, including ambulation, hindlimb foot angle, surface righting, negative geotaxis, front- and hindlimb suspension, grasping reflex, four limb grip strength and cliff aversion. Mice between the ages of post-natal day 2 to 14 can be used. In addition, these tests can be used for a wide range of neurological and neuromuscular pathologies, including cerebral palsy, hypoxic-ischemic encephalopathy, traumatic brain injury, spinal cord injury, neurodegenerative diseases, and neuromuscular disorders. These tests can also be used to determine the effects of pharmacological agents, as well as other types of therapeutic interventions. In this paper, motor deficits were evaluated in a novel neonatal mouse model of cerebral palsy that combines hypoxia, ischemia and inflammation. Forty-eight hours after injury, five tests out of the nine showed significant motor deficits: ambulation, hindlimb angle, hindlimb suspension, four limb grip strength, and grasping reflex. These tests revealed weakness in the hindlimbs, as well as fine motor skills such as grasping, which are similar to the motor deficits seen in human cerebral palsy patients.

Presented is a concise battery of mouse neonatal motor tests. Using these tests, neonatal motor deficits can be demonstrated in a variety of neonatal motor disorders. By having a standardized set of tests, results from different studies can be compared, allowing for better and accurate reporting between groups.

As the sheer number of transgenic mice strains grow and rodent models of pediatric disease increase, there is an expanding need for a comprehensive, ...

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 ...

Regenerative Therapy by Suprachoroidal Cell Autograft in Dry Age-related Macular Degeneration: Preliminary In Vivo Report

This study is aimed at examining whether a suprachoroidal graft of autologous cells can improve best corrected visual acuity (BCVA) and responses to microperimetry (MY) in eyes affected by dry Age-related Macular Degeneration (AMD) over time through the production and secretion of growth factors (GFs) on surrounding tissue. Patients were randomly assigned to each study group. All patients were diagnosed with dry AMD and with BCVA equal to or greater than 1 logarithm of the minimum angle of resolution (logMAR). A suprachoroidal autologous graft by Limoli Retinal Restoration Technique (LRRT) was carried out on group A, which included 11 eyes from 11 patients. The technique was performed by implanting adipocytes, adipose-derived stem cells obtained from the stromal vascular fraction, and platelets from platelet-rich plasma in the suprachoroidal space. Conversely, group B, including 14 eyes of 14 patients, was used as a control group. For each patient, diagnosis was verified by confocal scanning laser ophthalmoscope and spectral domain-optical coherence tomography (SD-OCT). In group A, BCVA improved by 0.581 to 0.504 at 90 days and to 0.376 logMAR at 180 days (+32.20%) postoperatively. Furthermore, MY test increased by 11.44 dB to 12.59 dB at 180 days. The different cell types grafted behind the choroid were able to ensure constant GF secretion in the choroidal flow. Consequently, the results indicate that visual acuity (VA) in the grafted group can increase more than in the control group after six months.

Regenerative Therapy by Suprachoroidal Cell Autograft in Dry Age-related Macular Degeneration: Preliminary In Vivo Report

The goal of this study is to assess whether the suprachoroidal graft of adipose-derived stem cells included in the stromal vascular fraction and platelets derived from platelet-rich plasma by the Limoli Retinal Restoration Technique can improve visual acuity and retinal sensitivity responses in eyes affected by dry age-related macular degeneration.

This study is aimed at examining whether a suprachoroidal graft of autologous cells can improve best corrected visual acuity (BCVA) and responses to ...

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.

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 ...