Gli autori riconoscono un sostegno finanziario da National Science Foundation (NSF) concedere CEE-0.540.832 intitolato &quot;Tecnologie Mid-Infrared per la Salute e l&#39;Ambiente (Mirthe)&quot;

1. Preparazione degli strumenti <ol><li> Accendere l&#39;alimentatore esterno per l&#39;ammoniaca piattaforma sensore ottico, un controller per diodo laser, una unità elettronica di controllo personalizzato costruito (CEU), il campionatore respiro, pompa di aria, e computer portatile. </li><li> Verificare che sia lo scarico e ventole di raffreddamento del sensore di ammoniaca sono in funzione. NOTA: Uno si trova nella parte posteriore del sensore, il secondo si trova all&#39;interno del sensore, che è facilmente accessibile. </li><li> Assicurano il modulo di rilevamento acustico e temperatura valvola a spillo sono a 38,0 ° C controllando il display digitale posizionato sul lato della scatola sensore ammoniaca. Attendere circa 35 minuti dal momento in cui il sensore è fornito di alimentazione per la temperatura si stabilizzi. </li><li> Set di tubi e temperatura di ingresso boccaglio a 55 ° C, cliccando sul nome di un icona &quot;ChangeT&quot; che si trova sul desktop del campionatore respiro. La temperatura può essere modificata facendo clic sulla freccia su o giù, seguito cliccando su &quot;Updmangiato Temp &quot;. Fare clic su &quot;Esci&quot; per tornare al desktop. Lasciare che il sistema almeno 5 minuti per la temperatura si stabilizzi. Mantenere la temperatura del campionatore e il tubo a 55 ° C minimizza la perdita superficie di ammoniaca. </li><li> Aprire il software sul computer portatile che controlla il sensore ammoniaca. Il programma può essere raggiunto all&#39;interno di una cartella denominata &quot;NH 3 Breath programma Sensor&quot; sul desktop. All&#39;interno di questa cartella, l&#39;utente deve selezionare l&#39;icona &quot;principali software labview&quot;. Questa cartella contiene diverse applicazioni, ma l&#39;utente deve selezionare &quot;Mainsequence.vi&quot; per accedere all&#39;interfaccia desiderata. Selezionare &quot;run&quot; nell&#39;angolo in alto a sinistra dello schermo. NOTA: Questo inizia la linea di blocco sequenza di taratura. Il laser del monitor QEPAS opera su una corrente ottimale, che viene selezionato durante una procedura di linea-bloccaggio automatico. Questo processo richiederà circa 25 min. </li><li> Creare una nuova sessione sul campionatore tale che EAch sessione soggetto ha un proprio file che viene salvato in un flash drive collegata. NOTA: Questa operazione viene eseguita aprendo il programma &quot;Respiro Sampler&quot; che si trova sul desktop del campionatore. C&#39;è spazio per identificare la sessione appropriato. Tutti i dati generati durante la sessione verranno salvati nella flash drive sotto questo identificativo. La data dell&#39;esperimento viene in genere utilizzato come parte del nome del file. L&#39;ingresso e temperatura boccaglio devono essere regolati prima di entrare nel programma respiro-campionamento. </li><li> Inserire un nuovo boccaglio monouso nel tubo di aspirazione. Indossare guanti monouso per evitare di contaminare boccaglio con ammoniaca dalle dita. </li></ol> 2. Breath Raccolta dei campioni NOTA: La competente Institutional Review Board (Comitato Etico) deve approvare qualsiasi studio che coinvolge soggetti umani. Ci sono molti fattori che possono influenzare drasticamente respiro ammoniaca. Questi fattori possono alterare respiro misurazioni ammoniaca direttamente da unffecting livelli di ammoniaca sistemici o modificando il viaggio di metaboliti respiro dai polmoni alla strumentazione. <ol><li> Assicurarsi che i soggetti arrivano al laboratorio in uno stato di digiuno, dopo aver consumato cibo per circa 12 ore all&#39;arrivo e che hanno evitato di esercizio la mattina prima del test. </li><li> Assicurarsi che nessuna sostanza è stato introdotto in bocca per almeno 1 ora prima della raccolta dei dati. Assicurarsi che i soggetti lavano i denti superiori a 1 ora prima del test. </li><li> Sedile oggetto davanti al sensore ammoniaca. Istruire il soggetto a tenere il tubo di aspirazione, e assicurarsi che non tocchino il boccaglio per evitare la contaminazione di ammoniaca. </li><li> Fare clic su &quot;Start&quot; sull&#39;interfaccia campionatore. Hanno il soggetto espirare nel boccaglio a lungo possibile, o fino a quando l&#39;operatore ritiene il campione sia sufficiente. NOTA: Questo è un singolo espirazione completa della durata di almeno 10 sec. Pressione bocca viene misurata in tempo reale come un surrogatoper la portata. Un manometro codice colore aiuta i prodotti tematici e mantenere la pressione di espirazione desiderata di 10 cm di acqua, con un tasso di flusso di espirazione di 50 ml / sec. Questo flusso è stato scelto in quanto è stato adottato dal ATS / ERS per il protocollo per determinare Fe NO. Questo tasso di flusso espiratorio è realizzabile da bambini e adulti. Allo stesso modo tre esalazioni riproducibili devono essere ottenuti che differiscono di meno del 10%. </li><li> Fare clic su &quot;Stop&quot; sull&#39;interfaccia campionatore quando il campione di aria espirata è completata. </li></ol> 3. Campione di respiro misura <ol><li> Una volta che un campione di aria espirata è stato analizzato, il tecnico di laboratorio funzionare il monitor ha quindi la capacità di analizzare qualsiasi segmento di tale profilo respiro. La porzione del respiro che è di interesse è il segmento di fase III. Questa è caratterizzata da un &quot;plateau&quot; della concentrazione di anidride carbonica e si trova al centro di fase tardiva della breath. </li><li> Selezionare la fase III porzione del campione trascinando le linee verticali sull&#39;interfaccia campionatore per iniziare quando altipiani di anidride carbonica e fermare subito prima della caduta di pressione del respiro. Vedere la Figura 1 per chiarimenti. </li><li> Salvare i dati sul flash drive cliccando su &quot;Store&quot; sull&#39;interfaccia campionatore touch screen. </li><li> Una volta che i dati sono stati memorizzati respiro, l&#39;utente può selezionare &quot;start&quot; per avviare un nuovo campione di respiro. </li></ol> 4. Illustra gli effetti del Collutorio e del pH sulla Breath Ammoniaca <ol><li> Campione 3 respiri per stabilire il livello di ammoniaca di base. Assicurarsi che respiri vanno prelevati almeno 5 minuti l&#39;uno dall&#39;altro. </li><li> Sciacquare accuratamente la bocca con un&#39;aliquota di 30 ml di acqua per 60 sec. </li><li> Raccogliere un campione di respiro entro 60 secondi del risciacquo (sezione 2). Raccogliere campioni di aria espirata nel corso della prossima ora ad osservare la variazione di ammoniaca nel tempo. NOTA: Samples può essere preso come frequentemente come ogni minuto, ma intervalli più lunghi sono tipicamente utilizzati. </li><li> Sciacquare la bocca con una aliquota di 30 ml di soluzione di base (bicarbonato di sodio in acqua) per 60 sec e ripetere 4.2.1. </li><li> Sciacquare la bocca accuratamente con una aliquota di 30 ml di soluzione acida per 60 secondi e ripetere 4.2.1. </li></ol> 5. Illustra gli effetti del Inlet e dei Trasporti tubi Temperature in Breath Ammoniaca <ol><li> Esempi tre respiri nel corso di 15 min con la temperatura di ingresso impostati sotto della temperatura corporea, circa 30 ° C. </li><li> Aumentare le temperature di ingresso e tubi di trasporto a 55 ° C, rispettivamente con una icona sul desktop sull&#39;interfaccia touch screen del campionatore respiro. Lasciare che il sistema almeno 5 min per raggiungere lo stato stazionario. </li><li> Esempio 3 respiri, 5 min a parte nella presa riscaldata. </li></ol>

<ol>
	<li>Adeva, M. M., Souto, G., Blanco, N., Donapetry, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ammonium+metabolism+in+humans.">Ammonium metabolism in humans.</a> Metabolism: clinical and experimental. 61 (11), 1495-1511 (2012).</li><li>Auron, A., Brophy, P. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hyperammonemia+in+review:+pathophysiology,+diagnosis,+and+treatment.">Hyperammonemia in review: pathophysiology, diagnosis, and treatment.</a> Pediatric nephrology. 27 (2), 207-222 (2012).</li><li>Blanco Vela, C. I., Bosques Padilla, F. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Determination+of+ammonia+concentrations+in+cirrhosis+patients-still+confusing+after+all+these+years.">Determination of ammonia concentrations in cirrhosis patients-still confusing after all these years.</a> Annals of hepatology. 10 Suppl 2, (2011).</li><li>Mpabanzi, L., Ol de Damink, S. W. M., van de Poll, M. C. G., Soeters, P. B., Jalan, R., Dejong, C. H. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=To+pee+or+not+to+pee:+ammonia+hypothesis+of+hepatic+encephalopathy+revisited.">To pee or not to pee: ammonia hypothesis of hepatic encephalopathy revisited.</a> European journal of gastroenterology &amp; hepatology. 23 (6), 449-454 (2011).</li><li>Goggs, R., Serrano, S., Szladovits, B., Keir, I., Ong, R., Hughes, 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+investigation+of+a+point-of-care+blood+ammonia+analyzer.">Clinical investigation of a point-of-care blood ammonia analyzer.</a> Veterinary clinical pathology / American Society for Veterinary Clinical Pathology. 37 (2), 198-206 (2008).</li><li>Huizenga, J. R., Tangerman, A., Gips, C. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Determination+of+ammonia+in+biological+fluids.">Determination of ammonia in biological fluids.</a> Annals of clinical biochemistry. 31 (Pt 6), 529-543 (1994).</li><li>Lewicki, 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=Real+time+ammonia+detection+in+exhaled+human+breath+with+a+quantum+cascade+laser+based+sensor.">Real time ammonia detection in exhaled human breath with a quantum cascade laser based sensor.</a> 2009 Conference on Lasers and ElectroOptics and 2009 Conference on Quantum electronics and Laser Science Conference. 1, (2009).</li><li>. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=American+Thoracic+Society.+European+Respiratory+Society.+Recommendations+for+Standardized+Procedures+for+the+Online+and+Offline+Measurement+of+Exhaled+Lower+Respiratory+Nitric+Oxide+and+Nasal+Nitric+Oxide.">American Thoracic Society. European Respiratory Society. Recommendations for Standardized Procedures for the Online and Offline Measurement of Exhaled Lower Respiratory Nitric Oxide and Nasal Nitric Oxide.</a> American journal of respiratory and critical care medicine. 171 (8), 912-930 (2005).</li><li>Solga, S. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Factors+influencing+breath+ammonia+determination.">Factors influencing breath ammonia determination.</a> Journal of breath research. 7 (3), (2013).</li><li>Schmidt, F. 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=Ammonia+in+breath+and+emitted+from+skin.">Ammonia in breath and emitted from skin.</a> Journal of breath research. 7 (1), (2013).</li><li>Span&#283;l, P., Dryahina, K., Smith, 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+quantitative+study+of+the+influence+of+inhaled+compounds+on+their+concentrations+in+exhaled+breath.">A quantitative study of the influence of inhaled compounds on their concentrations in exhaled breath.</a> Journal of breath research. 7 (1), (2013).</li><li>Boots, A. W., van Berkel, J. J. B. N., Dallinga, J. W., Smolinska, A., Wouters, E. F., van Schooten, F. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+versatile+use+of+exhaled+volatile+organic+compounds+in+human+health+and+disease.">The versatile use of exhaled volatile organic compounds in human health and disease.</a> Journal of breath research. 6 (2), (2012).</li><li>Hibbard, T., Killard, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Breath+ammonia+levels+in+a+normal+human+population+study+as+determined+by+photoacoustic+laser+spectroscopy.">Breath ammonia levels in a normal human population study as determined by photoacoustic laser spectroscopy.</a> Journal of breath research. 5 (3), (2011).</li><li>Wang, T., Pysanenko, A., Dryahina, K., Span&#283;l, P., Smith, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analysis+of+breath,+exhaled+via+the+mouth+and+nose,+and+the+air+in+the+oral+cavity.">Analysis of breath, exhaled via the mouth and nose, and the air in the oral cavity.</a> Journal of breath research. 2 (3), (2008).</li><li>Amann, A., Smith, 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> Volatile Biomarkers. 1st Edition. , (2013).</li></ol>

Dr. Risby serves as a consultant for Loccioni Humancare the manufacturer of the breath sampler.

I vantaggi di una procedura non invasiva in grado di rilevare metaboliti traccia in tempo reale sono evidenti. Tuttavia, il campo della ricerca fiato ha lottato per adempiere a tale potenziale. Misurazione Breath è un processo dinamico vulnerabile a molti fattori confondenti. Il nostro approccio ha importanti punti di forza: in particolare, la sensibilità e la velocità del monitor ammoniaca basato Rice QEPAS accoppiato al campionatore respiro ci hanno permesso di valutare ed identificare respiro fattori raccolta germ...

L&#39;ammoniaca è un sottoprodotto ubiquitario della proteina metabolismo 1. Misurazione ammoniaca può quindi aiutare i medici a valutare varie malattie e stati di benessere 2. Tuttavia, l&#39;ammoniaca è difficile misurare con precisione, attraverso il sangue o soffio, perché è molto reattiva. Anche se comunemente usato, analisi del sangue hanno numerosi inconvenienti, tra cui preoccupazioni fondamentali circa la precisione 3. Ma il problema principale con analisi del sangue è la realtà che sempre e solo episodicamente raccolti. Questo è importante perché ammoniaca fisiologia, molto simile glicemia e molti altri processi metabolici, sono fluide e in continuo cambiamento 4. Al contrario, le analisi del respiro sono completamente non invasivo e rapido, quindi facilmente consentendo misure ripetute. Pertanto, la misurazione ammoniaca respiro è interessante perché può rivolgere un serio bisogno insoddisfatto in un modo unico. Raccolta Breath, tuttavia, presenta problemi particolari. Considerando che il salasso porta intrinsecamente il JEOPardy di errori in diversi modi imprevedibili (ad esempio, il tempo laccio emostatico, la contaminazione sudore, emolisi dei globuli, ritardo nella misura di laboratorio, ecc 5), i ricercatori di misurazione respiro deve vedersela con un gruppo diverso di nuove sfide: la variabilità nella respirazione, la contaminazione con mucosa orale o ammoniaca batterica, influenza dell&#39;aria ambiente ed apparecchi umidità e temperatura, ecc 6. Infatti, non è saggio sottovalutare il compito di collegare dispositivi sperimentali per gli esseri umani utilizzando procedure sperimentali per scoprire sconosciuti biologia. In parte a causa di questi ostacoli, il respiro di ammoniaca non ha ancora raggiunto il suo potenziale. Qui, vi presentiamo il nostro protocollo di misura ammoniaca respiro per risultati veloci e accurati. Il nostro protocollo ha la forza in tre aree: il monitor, il campionatore interfaccia, e l&#39;attenzione alle influenze umane. Il monitor è stato costruito dai colleghi della Rice University come descritto in precedenza 7. La base del MEAsurement è una spettroscopia fotoacustica rafforzata quarzo (QEPAS) tecnica che impiega un diapason quarzo piezoelettrico come un trasduttore acustico. Effetto fotoacustico si verifica quando le onde acustiche sono prodotte dall&#39;assorbimento di radiazione laser modulata da gas in tracce bersaglio. Il gas di traccia viene rilevato utilizzando una cella acustico che è acusticamente risonante alla frequenza modulata. Lunghezza d&#39;onda di assorbimento per l&#39;ammoniaca è stata selezionata che è libero da interferenze spettrali da interferire specie respiro. Ai fini della misurazione respiro esalato umano, le principali caratteristiche del monitor comprendono una vasta gamma di misura (da circa 50 parti per miliardo, ppb di almeno 5.000 ppb) e velocità (1 misurazioni sec). La velocità del monitor ha la risoluzione temporale durante il ciclo respiratorio. Il monitor è accoppiato ad un campionatore respiro appositamente progettato. Il campionatore è costituito da un sensore di pressione e capnografo. Esso mostra e archivia in tempo realemisure di pressione bocca e anidride carbonica, nonché le concentrazioni di ammoniaca determinata dal sensore. Questo campionatore, dunque, consente al tecnico di valutare la qualità dello sforzo respiro come il respiro è raccolto. Questo ci permette di superare le raccomandazioni per analizzare il respiro di ossido nitrico (NO Fe) proposto dalla Task Force della American Thoracic Society / European Respiratory Society (ATS / ERS) 8. Per tutti campionamento di aria espirata, una valvola unidirezionale monouso in linea è stato usato sulla porta bocca del campionatore respiro. A causa della velocità del monitor e dei controlli di qualità fornito dal campionatore, siamo stati in grado di valutare con attenzione le influenze umane 9. La maggior parte dei soggetti, per esempio, inizialmente iperventilazione quando richiesto per respirare. Altre influenze importanti, come il pH orale e collutori, temperature del campionatore, monitor e tutti i tubi associati, e le modalità di respirazione, sono stati poi studiati, e sono la base for gli esperimenti riportati qui di seguito. Infine, e forse più significativamente, si deve sottolineare che più gruppi di grande esperienza stanno misurando respiro ammoniaca utilizzando completamente diversi sensori e procedure di misurazione. Queste possono avere importanti vantaggi e validità. Un confronto completo è oltre la portata del presente 10,11,12 lavoro.

<table border="0" cellpadding="0" cellspacing="0" width="666">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:211px;">Rice Ammonia Monitor System</td>
			<td style="width:171px;">N/A</td>
			<td style="width:119px;">N/A</td>
			<td style="width:167px;">Not available for commercial purchase</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:211px;">Loccioni Breath Sampler</td>
			<td style="width:171px;">Loccioni Humancare</td>
			<td style="width:119px;">N/A</td>
			<td>Single breath version</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:211px;">Disposable Mouth Piece</td>
			<td style="width:171px;">WestPrime Healthcare</td>
			<td style="width:119px;">G011-200</td>
			<td>Manufacturer is AlcoQuant</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:211px;">Laptop</td>
			<td style="width:171px;">Lenovo</td>
			<td style="width:119px;">N/A</td>
			<td>Old model no longer sold by manufacturer</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:211px;">Acid Rinse</td>
			<td style="width:171px;">N/A</td>
			<td style="width:119px;">N/A</td>
			<td>Household acidic drink (coffee, soft drink, citrus juices, etc)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:211px;">Base Rinse</td>
			<td style="width:171px;">N/A</td>
			<td style="width:119px;">N/A</td>
			<td>Water mixed with a nonexact amount of sodium bicarbonate (Arm &#38; Hammer Baking Soda)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:211px;">Neutral Rinse</td>
			<td style="width:171px;">N/A</td>
			<td style="width:119px;">N/A</td>
			<td>Water</td>
		</tr>
	</tbody>
</table>

fast and accurate exhaled breath ammonia measurement

This exhaled breath ammonia method uses a fast and highly sensitive spectroscopic method known as quartz enhanced photoacoustic spectroscopy (QEPAS) that uses a quantum cascade based laser. The monitor is coupled to a sampler that measures mouth pressure and carbon dioxide. The system is temperature controlled and specifically designed to address the reactivity of this compound. The sampler provides immediate feedback to the subject and the technician on the quality of the breath effort. Together with the quick response time of the monitor, this system is capable of accurately measuring exhaled breath ammonia representative of deep lung systemic levels. 
Because the system is easy to use and produces real time results, it has enabled experiments to identify factors that influence measurements. For example, mouth rinse and oral pH reproducibly and significantly affect results and therefore must be controlled. Temperature and mode of breathing are other examples. As our understanding of these factors evolves, error is reduced, and clinical studies become more meaningful. This system is very reliable and individual measurements are inexpensive. 
The sampler is relatively inexpensive and quite portable, but the monitor is neither. This limits options for some clinical studies and provides rational for future innovations.

I soggetti possono essere tenuti a produrre una vasta gamma di livelli basali di ammoniaca respiro. Individui sani possono iniziare la giornata con una misura di ammoniaca soffio di 100-1.000 ppb. Sciacquare la bocca con il liquido cambia immediatamente l&#39;importo delle rilevabile respiro ammoniaca. Liquidi neutri e acidi in genere tagliare la quantità di ammoniaca osservabile di oltre la metà. Questi livelli poi ritornano ai valori basali come gli effetti del risciacquo svanisce. Gli effetti di acqua sembrano diss...

Watch this Scientific Journal Video about Fast and Accurate Exhaled Breath Ammonia Measurement at JoVE.com

Fast and Accurate Exhaled Breath Ammonia Measurement

Ammonia is an important physiologic metabolite relevant to various disease and wellness states. It is also a difficult molecule to measure in breath, which demands particular precautions be taken to obtain accurate results. Not all factors influencing ammonia are known, but progress can be difficult without accounting for these factors.

Veloce e preciso espirata Ammoniaca misura

This exhaled breath ammonia method uses a fast and highly sensitive spectroscopic method known as quartz enhanced photoacoustic spectroscopy (QEPAS) that ...

fast-and-accurate-exhaled-breath-ammonia-measurement

Research

JoVE Journal

Medicine

13.1K Views.  St. Luke's University Hospital. Ammonia is an important physiologic metabolite relevant to various disease and wellness states. It is also a difficult molecule to measure in breath, which demands particular precautions be taken to obtain accurate results. Not all factors influencing ammonia are known, but progress can be difficult without accounting for these factors.

Video: Fast and Accurate Exhaled Breath Ammonia Measurement

Accurate and Simple Measurement of the Pro-inflammatory Cytokine IL-1&beta; using a Whole Blood Stimulation Assay

Inflammatory processes resulting from the secretion of soluble mediators by immune cells, lead to various manifestations in skin, joints and other tissues as well as altered cytokine homeostasis. The innate immune system plays a crucial role in recognizing pathogens and other endogenous danger stimuli. One of the major cytokines released by innate immune cells is Interleukin (IL)-1. Therefore, we utilize a whole blood stimulation assay in order to measure the secretion of inflammatory cytokines and specifically of the pro-inflammatory cytokine IL-1&beta; 1, 2, 3.
Patients with genetic dysfunctions of the innate immune system causing autoinflammatory syndromes show an exaggerated release of mature IL-1&beta; upon stimulation with LPS alone. In order to evaluate the innate immune component of patients who present with inflammatory-associated pathologies, we use a specific immunoassay to detect cellular immune responses to pathogen-associated molecular patterns (PAMPs), such as the gram-negative bacterial endotoxin, lipopolysaccharide (LPS). These PAMPs are recognized by pathogen recognition receptors (PRRs), which are found on the cells of the innate immune system 4, 5, 6, 7. A primary signal, LPS, in conjunction with a secondary signal, ATP, is necessary for the activation of the inflammasome, a multiprotein complex that processes pro-IL-1&beta; to its mature, bioactive form 4, 5, 6, 8, 9, 10.
The whole blood assay requires minimal sample manipulation to assess cytokine production when compared to other methods that require labor intensive isolation and culturing of specific cell populations. This method differs from other whole blood stimulation assays; rather than diluting samples with a ratio of RPMI media, we perform a white blood cell count directly from diluted whole blood and therefore, stimulate a known number of white blood cells in culture 2. The results of this particular whole blood assay demonstrate a novel technique useful in elucidating patient cohorts presenting with autoinflammatory pathophysiologies.

Accurate and Simple Measurement of the Pro-inflammatory Cytokine IL-1&amp;#946; using a Whole Blood Stimulation Assay

We describe a simple immunoassay to measure the production of pro-inflammatory cytokines, such as IL-1 beta production, in patients presenting with autoinflammatory phenotypes. By activating cells in whole blood cultures with pathogen-associated molecular patterns, specifically with lipopolysaccharide, cytokine secretion can be conveniently evaluated in whole blood supernatants.

Misurazione accurata e semplice della citochina pro-infiammatoria IL-1β utilizzando un intero saggio stimolazione del sangue

Inflammatory processes resulting from the secretion of soluble mediators by immune cells, lead to various manifestations in skin, joints and other tissues ...

Ricerca

Medicina

Accurate and Simple Evaluation of Vascular Anastomoses in Monochorionic Placenta using Colored Dye

The presence of placental vascular anastomoses is a conditio sine qua non for the development of twin-to-twin transfusion syndrome (TTTS) and twin anemia polycythemia sequence (TAPS)1,2. Injection studies of twin placentas have shown that such anastomoses are almost invariably present in monochorionic twins and extremely rare in dichorionic twins1. Three types of anastomoses have been documented: from artery to artery, from vein to vein and from artery to vein. Arterio-venous (AV) anastomoses are unidirectional and are referred to as "deep" anastomoses since they proceed through a shared placental cotyledon, whereas arterio-arterial (AA) and veno-venous (VV) anastomoses are bi-directional and are referred to as "superficial" since they lie on the chorionic plate. Both TTTS and TAPS are caused by net imbalance of blood flow between the twins due to AV anastomoses. Blood from one twin (the donor) is pumped through an artery into the shared placental cotyledon and then drained through a vein into the circulation of the other twin (the recipient). Unless blood is pumped back from the recipient to the donor through oppositely directed deep AV anastomoses or through superficial anastomoses, an imbalance of blood volumes occurs, gradually leading to the development of TTTS or TAPS. The presence of an AA anastomosis has been shown to protect against the development of TTTS and TAPS by compensating for the circulatory imbalance caused by the uni-directional AV anastomoses1,2. 
Injection of monochorionic placentas soon after birth is a useful mean to understand the etiology of various (hematological) complications in monochorionic twins and is a required test to reach the diagnosis of TAPS2. In addition, injection of TTTS placentas treated with fetoscopic laser surgery allows identification of possible residual anastomoses3-5. This additional information is of paramount importance for all perinatologists involved in the management and care of monochorionic twins with TTTS or TAPS. Several placental injection techniques are currently being used. We provide a simple protocol to accurately evaluate the presence of (residual) vascular anastomoses using colored dye injection.

Twin-to-twin transfusion syndrome and twin anemia polycythemia sequence are two potentially devastating problems in perinatal medicine. Both disorders occur only in monochorionic twins and result from unbalanced blood flow through placental vascular anastomoses. We provide a simple protocol to accurately evaluate the presence of vascular anastomoses using colored dye injection of placental vessels after birth.

La valutazione accurata e semplice delle anastomosi vascolari in placenta monocoriale con colorante

The presence of placental vascular anastomoses is a conditio sine qua non for the development of twin-to-twin transfusion syndrome (TTTS) and twin anemia ...

Remote Magnetic Navigation for Accurate, Real-time Catheter Positioning and Ablation in Cardiac Electrophysiology Procedures

New remote navigation systems have been developed to improve current limitations of conventional manually guided catheter ablation in complex cardiac substrates such as left atrial flutter. This protocol describes all the clinical and invasive interventional steps performed during a human electrophysiological study and ablation to assess the accuracy, safety and real-time navigation of the Catheter Guidance, Control and Imaging (CGCI) system. Patients who underwent ablation of a right or left atrium flutter substrate were included. Specifically, data from three left atrial flutter and two counterclockwise right atrial flutter procedures are shown in this report. One representative left atrial flutter procedure is shown in the movie. This system is based on eight coil-core electromagnets, which generate a dynamic magnetic field focused on the heart. Remote navigation by rapid changes (msec) in the magnetic field magnitude and a very flexible magnetized catheter allow real-time closed-loop integration and accurate, stable positioning and ablation of the arrhythmogenic substrate.

This report provides a detailed description of a new remote navigation system based on magnetic driven forces, which has been recently introduced as a new robotic tool for human cardiac electrophysiology procedures.

Navigazione magnetica a distanza per Preciso, posizionamento del catetere in tempo reale e ablazione in Elettrofisiologia Cardiaca Procedure

New remote navigation systems have been developed to improve current limitations of conventional manually guided catheter ablation in complex cardiac ...

Assessment of Gastric Emptying in Non-obese Diabetic Mice Using a [13C]-octanoic Acid Breath Test

Gastric emptying studies in mice have been limited by the inability to follow gastric emptying changes in the same animal since the most commonly used techniques require killing of the animals and postmortem recovery of the meal1,2. This approach prevents longitudinal studies to determine changes in gastric emptying with age and progression of disease. The commonly used [13C]-octanoic acid breath test for humans3 has been modified for use in mice4-6 and rats7 and we previously showed that this test is reliable and responsive to changes in gastric emptying in response to drugs and during diabetic disease progression8. In this video presentation the principle and practical implementation of this modified test is explained. As in the previous study, NOD LtJ mice are used, a model of type 1 diabetes9. A proportion of these mice develop the symptoms of gastroparesis, a complication of diabetes characterized by delayed gastric emptying without mechanical obstruction of the stomach10.
This paper demonstrates how to train the mice for testing, how to prepare the test meal and obtain 4 hr gastric emptying data and how to analyze the obtained data. The carbon isotope analyzer used in the present study is suitable for the automatic sampling of the air samples from up to 12 mice at the same time. This technique allows the longitudinal follow-up of gastric emptying from larger groups of mice with diabetes or other long-standing diseases.

Assessment of Gastric Emptying in Non-obese Diabetic Mice Using a [13C]-octanoic Acid Breath Test

Determination of gastric emptying with a non-invasive [13C]-octanoic acid breath test for tracking gastroparesis in female NOD LtJ mice.

La valutazione di svuotamento gastrico in non-obesi topi diabetici utilizzando un [<sup&gt; 13</sup&gt; C]-ottanoico Breath Test Acid

Gastric emptying studies in mice have been limited by the inability to follow gastric emptying changes in the same animal since the most commonly used ...

Lesion Explorer: A Video-guided, Standardized Protocol for Accurate and Reliable MRI-derived Volumetrics in Alzheimer's Disease and Normal Elderly

Obtaining in vivo human brain tissue volumetrics from MRI is often complicated by various technical and biological issues. These challenges are exacerbated when significant brain atrophy and age-related white matter changes (e.g. Leukoaraiosis) are present. Lesion Explorer (LE) is an accurate and reliable neuroimaging pipeline specifically developed to address such issues commonly observed on MRI of Alzheimer's disease and normal elderly. The pipeline is a complex set of semi-automatic procedures which has been previously validated in a series of internal and external reliability tests1,2. However, LE's accuracy and reliability is highly dependent on properly trained manual operators to execute commands, identify distinct anatomical landmarks, and manually edit/verify various computer-generated segmentation outputs.
LE can be divided into 3 main components, each requiring a set of commands and manual operations: 1) Brain-Sizer, 2) SABRE, and 3) Lesion-Seg. Brain-Sizer's manual operations involve editing of the automatic skull-stripped total intracranial vault (TIV) extraction mask, designation of ventricular cerebrospinal fluid (vCSF), and removal of subtentorial structures. The SABRE component requires checking of image alignment along the anterior and posterior commissure (ACPC) plane, and identification of several anatomical landmarks required for regional parcellation. Finally, the Lesion-Seg component involves manual checking of the automatic lesion segmentation of subcortical hyperintensities (SH) for false positive errors.
While on-site training of the LE pipeline is preferable, readily available visual teaching tools with interactive training images are a viable alternative. Developed to ensure a high degree of accuracy and reliability, the following is a step-by-step, video-guided, standardized protocol for LE's manual procedures.

Lesion Explorer: A Video-guided, Standardized Protocol for Accurate and Reliable MRI-derived Volumetrics in Alzheimer&#039;s Disease and Normal Elderly

Lesion Explorer (LE) is a semi-automatic, image-processing pipeline developed to obtain regional brain tissue and subcortical hyperintensity lesion volumetrics from structural MRI of Alzheimer's disease and normal elderly. To ensure a high level of accuracy and reliability, the following is a video-guided, standardized protocol for LE's manual procedures.

Lesione Explorer: un video-guida, standardizzato protocollo preciso e affidabile Volumetrics MRI-derivati ​​nella malattia di Alzheimer e normale Anziani

Obtaining in vivo human brain tissue volumetrics from MRI is often complicated by various technical and biological issues. These challenges are ...

Voluntary Breath-hold Technique for Reducing Heart Dose in Left Breast Radiotherapy

Breath-holding techniques reduce the amount of radiation received by cardiac structures during tangential-field left breast radiotherapy. With these techniques, patients hold their breath while radiotherapy is delivered, pushing the heart down and away from the radiotherapy field. Despite clear dosimetric benefits, these techniques are not yet in widespread use. One reason for this is that commercially available solutions require specialist equipment, necessitating not only significant capital investment, but often also incurring ongoing costs such as a need for daily disposable mouthpieces. The voluntary breath-hold technique described here does not require any additional specialist equipment. All breath-holding techniques require a surrogate to monitor breath-hold consistency and whether breath-hold is maintained. Voluntary breath-hold uses the distance moved by the anterior and lateral reference marks (tattoos) away from the treatment room lasers in breath-hold to monitor consistency at CT-planning and treatment setup. Light fields are then used to monitor breath-hold consistency prior to and during radiotherapy delivery.

The current priority in breast cancer radiotherapy is to reduce cardiac doses without compromising target tissue coverage. The voluntary breath-hold technique described here is a simple, inexpensive solution to this problem and capable of being instituted widely without the need for specialized equipment.

Volontariato Tecnica Breath-hold per la riduzione Cuore Dose in Radioterapia della mammella sinistra

Breath-holding techniques reduce the amount of radiation received by cardiac structures during tangential-field left breast radiotherapy. With these ...

Creating Anatomically Accurate and Reproducible Intracranial Xenografts of Human Brain Tumors

Orthotopic tumor models are currently the best way to study the characteristics of a tumor type, with and without intervention, in the context of a live animal &#8211;&#160;particularly in sites with unique physiological and architectural qualities such as the brain. In vitro and ectopic models cannot account for features such as vasculature, blood brain barrier, metabolism, drug delivery and toxicity, and a host of other relevant factors. Orthotopic models have their limitations too, but with proper technique tumor cells of interest can be accurately engrafted into tissue that most closely mimics conditions in the human brain. By employing methods that deliver precisely measured volumes to accurately defined locations at a consistent rate and pressure, mouse models of human brain tumors with predictable growth rates can be reproducibly created and are suitable for reliable analysis of various interventions. The protocol described here focuses on the technical details of designing and preparing for an intracranial injection, performing the surgery, and ensuring successful and reproducible tumor growth and provides starting points for a variety of conditions that can be customized for a range of different brain tumor models.

The brain is a unique site with qualities that are not well represented by in vitro or ectopic analyses. Orthotopic mouse models with reproducible location and growth characteristics can be reliably created with intracranial injections using a stereotaxic fixation instrument and a low pressure syringe pump.

Creazione anatomicamente accurati e riproducibili intracranica xenotrapianti di tumori cerebrali umani

Orthotopic tumor models are currently the best way to study the characteristics of a tumor type, with and without intervention, in the context of a live ...

3D Ultrasound Imaging: Fast and Cost-effective Morphometry of Musculoskeletal Tissue

The developmental goal of 3D ultrasound imaging (3DUS) is to engineer a modality to perform 3D morphological ultrasound analysis of human muscles. 3DUS images are constructed from calibrated freehand 2D B-mode ultrasound images, which are positioned into a voxel array. Ultrasound (US) imaging allows quantification of muscle size, fascicle length, and angle of pennation. These morphological variables are important determinants of muscle force and length range of force exertion. The presented protocol describes an approach to determine volume and fascicle length of m.&#160;vastus lateralis and m.&#160;gastrocnemius medialis. 3DUS facilitates standardization using 3D anatomical references. This approach provides a fast and cost-effective approach for quantifying 3D morphology in skeletal muscles. In healthcare and sports, information on the morphometry of muscles is very valuable in diagnostics and/or follow-up evaluations after treatment or training.

3D ultrasound imaging (3DUS) allows fast and cost-effective morphometry of musculoskeletal tissues. We present a protocol to measure muscle volume and fascicle length using 3DUS.

Ecografia 3D: veloce e conveniente morfometria del tessuto muscolo-scheletrico

The developmental goal of 3D ultrasound imaging (3DUS) is to engineer a modality to perform 3D morphological ultrasound analysis of human muscles. 3DUS ...

Fast Inspection of Quality of Indigo Naturalis by Multiple Light Scattering

Quality control of Chinese herbal medicine is a crucial component of Chinese herbal medicine research and development. Faced with the challenges of modernization and internationalization of Chinese herbal medicine, it is urgent to establish thorough and effective procedures for quality identification of Chinese herbal medicine, and there is an urgent need for new analytical and testing techniques that are efficient, accurate, and environmentally friendly. 
Multiple light scattering is a cutting-edge and analytical method that can accurately and rapidly assess the quality of Chinese herbal medicine without altering the nature or state of the sample or using organic reagents. Indigo Naturalis is considered a good remedy for pediatric hyperthermia, psoriasis, leukemia, and ulcerative colitis. In this study, the process of addition of Indigo Naturalis powder in water was recorded precisely using a multiple light scattering instrument. 
The qualitative and quantitative measurements of the instrument can be used to accurately capture the overall trajectory and sinking behavior of Indigo Naturalis powder into water and to establish a rapid evaluation method for the quality of Indigo Naturalis with the transmission and backscattering spectrograms of the sample as qualitative indicators and stability index as a quantitative indicator. The analytical technique based on multiple light scattering provides a fast, accurate, green, and environmentally friendly method for the quality evaluation of Indigo Naturalis and supports the development and transformation of high-quality Indigo Naturalis.

Here, we present a quality evaluation method based on a multiple light scattering technique to evaluate the quality of Indigo Naturalis.

Ispezione rapida della qualità di Indigo Naturalis mediante diffusione multipla della luce

Quality control of Chinese herbal medicine is a crucial component of Chinese herbal medicine research and development. Faced with the challenges of ...

Focused Assessment with Sonography for Trauma (FAST) Exam: Image Acquisition

Over the past twenty years, the Focused Assessment with Sonography for Trauma (FAST) exam has transformed the care of patients presenting with a combination of trauma (blunt or penetrating) and hypotension. In these hemodynamically unstable trauma patients, the FAST exam permits rapid and noninvasive screening for free pericardial or peritoneal fluid, the latter of which implicates intra-abdominal injury as a likely contributor to the hypotension and justifies emergent abdominal surgical exploration. Further, the abdominal portion of the FAST exam can also be used outside of the trauma setting to screen for free peritoneal fluid in patients who become hemodynamically unstable in any context, including after procedures that may inadvertently injure abdominal organs. These "non-trauma" situations of hemodynamic instability are often triaged by providers from specialties other than emergency medicine or trauma surgery who are not familiar with the FAST exam. Therefore, there is a need to promulgate knowledge about the FAST exam to all clinicians caring for critically ill patients. Toward this end, this article describes FAST exam image acquisition: patient positioning, transducer selection, image optimization, and exam limitations. Since the free fluid is likely to be found in specific anatomic locations that are unique for each canonical FAST exam view, this work centers on the unique image acquisition considerations for each window: subcostal, right upper quadrant, left upper quadrant, and pelvis.

Focused Assessment with Sonography for Trauma FAST Exam: Image Acquisition

The Focused Assessment with Sonography for Trauma (FAST) exam is a diagnostic point-of-care ultrasound examination used to screen for the presence of free fluid in the pericardium and peritoneum. Indications, techniques, and pitfalls of the procedure are discussed in this article.