دورية أكاديمية
Improving Immunoassay Performance with Cleavable Blocking of Microarrays.
| العنوان: | Improving Immunoassay Performance with Cleavable Blocking of Microarrays. |
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| المؤلفون: | Shlyapnikov YM; Institute of Theoretical and Experimental Biophysics Russian Academy of Sciences, Pushchino 142290, Russia., Malakhova EA; Institute of Theoretical and Experimental Biophysics Russian Academy of Sciences, Pushchino 142290, Russia., Shlyapnikova EA; Institute of Theoretical and Experimental Biophysics Russian Academy of Sciences, Pushchino 142290, Russia. |
| المصدر: | Analytical chemistry [Anal Chem] 2021 Jan 19; Vol. 93 (2), pp. 1126-1134. Date of Electronic Publication: 2020 Dec 11. |
| نوع المنشور: | Journal Article; Research Support, Non-U.S. Gov't |
| اللغة: | English |
| بيانات الدورية: | Publisher: American Chemical Society Country of Publication: United States NLM ID: 0370536 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1520-6882 (Electronic) Linking ISSN: 00032700 NLM ISO Abbreviation: Anal Chem Subsets: MEDLINE |
| أسماء مطبوعة: | Original Publication: Washington, American Chemical Society. |
| مواضيع طبية MeSH: | Fluorescent Antibody Technique* , Immunoassay* , Protein Array Analysis*, Cholera Toxin/*analysis , Cystine/*analogs & derivatives, Cholera Toxin/metabolism ; Cystine/chemical synthesis ; Cystine/chemistry ; Molecular Structure ; Vaccinia virus/enzymology ; Vaccinia virus/isolation & purification |
| مستخلص: | Among the key issues that are commonly associated with the development of microarray-based assays are nonspecific binding and diffusion constraints. Here we present a novel strategy addressing both of these challenges simultaneously. The essence of the method consists in blocking the microarray surface with a blocking agent containing a perfluoroalkyl chain and a disulfide linker. The resulting surface is hydrophobic, and no immiscible liquid layer remains on it upon cyclically draining and replenishing the sample solution, ensuring an efficient mass transfer of an analyte onto a microarray. Prior to the signal detection procedure, disulfide bonds are chemically cleaved, and the perfluoroalkyl chains are removed from the microarray surface along with nonspecifically adsorbed proteins, resulting in extremely low background. Using conventional fluorescent detection, we show a 30-fold increase in signal/background ratio compared to a common epoxy-modified glass substrate. The combination of this technique with magnetic beads detection results in a simple and ultrasensitive cholera toxin (CT) immunoassay. The limit of detection (LOD) is 1 fM, which is achieved with an analyte binding time of 1 h. Efficient mass transfer provides highly sensitive detection of whole virus particles despite their low diffusion coefficient. The achieved LOD for vaccinia virus is 10 4 particles in 1 mL of sample. Finally, we have performed for the first time the simultaneous detection of whole virus and CT protein biomarker in a single assay. The developed technique can be used for multiplex detection of trace amounts of pathogens of various natures. |
| المشرفين على المادة: | 48TCX9A1VT (Cystine) 9012-63-9 (Cholera Toxin) VEO23D129R (cystine dimethyl ester) |
| تواريخ الأحداث: | Date Created: 20201211 Date Completed: 20210218 Latest Revision: 20210218 |
| رمز التحديث: | 20240628 |
| DOI: | 10.1021/acs.analchem.0c04175 |
| PMID: | 33305941 |
| قاعدة البيانات: | MEDLINE |
Find this issue from the American Chemical Society | تدمد: | 1520-6882 |
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| DOI: | 10.1021/acs.analchem.0c04175 |