The designed benchtop technique is primed to detect bacteria and viruses from antigenic surface marker proteins in solutions, initially water. This inclusive bio-immunoassay uniquely combines nanofiltration and near infrared (NIR) dyes conjugated to antibodies to isolate and distinguish microbial antigens, using laser excitation and spectrometric analysis. The project goals include detecting microorganisms aboard the International Space Station, space shuttle, Crew Exploration Vehicle (CEV), and human habitats on future Moon and Mars missions, ensuring astronaut safety. The technique is intended to improve and advance water contamination testing both commercially and environmentally as well. Lastly, this streamlined technique poses to greatly simplify and expedite testing of pathogens in complex matrices, such as blood, in hospital and laboratory clinics.

The Benchtop Analysis System (a) consists of a medium-power, class IV laser, a four-port cuvette sampler holder, a spectrometer, laptop and software, and fiber-optic cables. (b) The microcentrifuge used in nanofiltration of the complexes.
The approach relies on NIR fluorescent dyes derivatized to specific antibody sets that are selected to bind and differentiate microbial surface proteins, termed antigens. In a solution containing an antigenic slurry, NIR conjugated antibodies are added to the mixture, and will bind to the respective antigens if present. To eliminate any false positives, excess antibodies, i.e., those antibodies not bound to an antigenic protein or those with no respective antigen present, are removed via the nanofiltration process using a portable, table-top centrifuge. The remaining NIR dye/ antibody and antigenic protein pairs left on the nanofilter are transferred to cuvette, excited by an NIR laser, and detected by spectrometer. Using simple computer software, the results are easily interpreted as intensity peaks at the appropriate NIR offset wavelength emission.

Initial data reveal the assay sensitively identified antigens at intensity counts of 100 IC or higher (or roughly 36 pW) with an accuracy of 85 percent for 2-hour incubations and 75 percent for 3-hour incubations. Interestingly, samples incubated for less time (2 hours vs 3 hours) produced an increased percentage of antigen detection. Further testing at incubation times such as 1 hour or lower could potentially increase positive predictability based on the study’s results. Also encouraging were negative control experiments with nonspecific antigens, beta galactosidase and thyroglobulin, which showed results of 100 percent accuracy, with no false positive detection.

This work was done by Maximilian C. Scardelletti and Vanessa Varaljay of Glenn Research Center.

Inquiries concerning rights for the commercial use of this invention should be addressed to

NASA Glenn Research Center
Innovative Partnerships Office
Attn: Steve Fedor
Mail Stop 4–8
21000 Brookpark Road
Cleveland
Ohio 44135.

Refer to LEW-18387-1.

Topics:
Bacteria Biological sciences Computer software and hardware Lasers Medical Optics Patient Monitoring Spacecraft Vehicles Water Water pollution Water quality Water treatment
This Brief includes a Technical Support Package (TSP).
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Benchtop Antigen Detection Technique Using Nanofiltration and Fluorescent Dyes

(reference LEW-18387-1) is currently available for download from the TSP library.

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Medical Design Briefs Magazine

This article first appeared in the May, 2009 issue of Medical Design Briefs Magazine (Vol. 33 No. 5).

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Overview

The document discusses a novel benchtop antigen detection technique developed by NASA's Glenn Research Center, which utilizes nanofiltration and fluorescent dyes to identify antigens in fluid samples. This innovation addresses the need for efficient, portable, and low-maintenance systems for detecting microbes and viruses, particularly in space missions, hospital laboratories, and bio-terrorism diagnostics.

Current detection methods, such as genetic assays and flow cytometry, while accurate, require bulky equipment, extensive laboratory facilities, and significant time for analysis. These methods often necessitate specialized personnel and can take days to yield results. In contrast, the new benchtop system aims to provide rapid and reliable detection of multiple microbes or viruses in a single sample, enhancing speed and portability.

The technique leverages near-infrared (NIR) fluorescent dyes, which improve detection accuracy by minimizing background noise due to low auto fluorescence in the NIR region. The detection process is based on the interaction between antibodies and antigens, where antibodies bind to specific sites on antigens, forming complexes that can be identified. The use of NIR dyes, such as IRDye™ 800, allows for specific detection of these complexes.

The document highlights the proof-of-concept experiments that demonstrate the effectiveness of this technique, achieving 100% accuracy with no false positives. The method involves centrifugation, nanofilters, and spectrometry, making it a comprehensive approach to antigen detection.

Overall, this benchtop antigen detection technique represents a significant advancement in microbiological diagnostics, offering a lightweight and efficient alternative to traditional methods. Its potential applications extend beyond space missions to include clinical settings and emergency response scenarios, where rapid detection of pathogens is critical. The development of this technology underscores NASA's commitment to leveraging aerospace-related innovations for broader scientific and commercial applications.