title: Ultraminiaturized assay for rapid, low cost detection and quantification of clinical and biochemical samples.
abstract: Herein, we report a simple, sensitive, rapid and low-cost ultraminiaturized assay technique for quantitative detection of 1 μl of clinical or biochemical sample on a novel ultraminiaturized assay plate (UAP). UAP is prepared by making tiny cavities on a polypropylene sheet. As UAP cannot immobilize a biomolecule through absorption, we have activated the tiny cavities of UAP by 1-fluoro-2-nitro-4-azidobenzene in a photochemical reaction. Activated UAP (AUAP) can covalently immobilize any biomolecule having an active nucleophilic group such as amino group. Efficacy of AUAP is demonstrated by detecting human IgE, antibody of hepatitis C virus core antigen and oligonucleotides. Quantification is performed by capturing the image of the colored assay solution and digitally quantifying the image by color saturation without using costly NanoDrop spectrophotometer. Image - based detection of human IgE and an oligonucleotide shows an excellent correlation with absorbance - based assay (recorded in a NanoDrop spectrophotometer); it is validated by Pearson's product-moment correlation with correlation coefficient of r = 0.9545088 and r = 0.9947444 respectively. AUAP is further checked by detecting hepatitis C virus Ab where strong correlation of color saturation with absorbance with respect to concentration is observed. Ultraminiaturized assay successfully detects target oligonucleotides by perfectly hybridizing with their respective complementary oligonucleotide probes but not with a random oligonucleotide. Ultraminiaturized assay technique has substantially reduced the requirement of reagents by 100 times and assay timing by 50 times making it a potential alternative to conventional method.
authors.affiliation: Innovative Diagnostic Lab, CSIR-Institute of Genomics and Integrative Biology, Mall Road, Delhi, 110 007, India.|Academy of Scientific and Innovative Research, CRRI Campus, Mathura Road, Delhi, 110020, India.
authors.forename: Shahila
authors.initials: S
authors.lastname: Parween
authors.affiliation: Innovative Diagnostic Lab, CSIR-Institute of Genomics and Integrative Biology, Mall Road, Delhi, 110 007, India. pnahar@igib.res.in.|Academy of Scientific and Innovative Research, CRRI Campus, Mathura Road, Delhi, 110020, India. pnahar@igib.res.in.
authors.forename: Pradip
authors.initials: P
authors.lastname: Nahar
mesh_terms: D000818:Animals; D000956:Antigens, Viral / Q000276:immunology; D001386:Azides / Q000737:chemistry; D003116:Color; D003365:Costs and Cost Analysis; D004797:Enzyme-Linked Immunosorbent Assay / Q000191:economics* / Q000379:methods*; D016174:Hepacivirus / Q000276:immunology; D006801:Humans; D007073:Immunoglobulin E / Q000032:analysis / Q000276:immunology; D008904:Miniaturization / Q000379:methods*; D009578:Nitrobenzenes / Q000737:chemistry; D009841:Oligonucleotides / Q000032:analysis; D055668:Photochemical Processes; D013997:Time Factors
chemical_list: C492759:1-fluoro-2-nitro-4-azidobenzene; D000956:Antigens, Viral; D001386:Azides; D009578:Nitrobenzenes; D009841:Oligonucleotides; D007073:Immunoglobulin E
keywords: Activated ultra miniaturized assay plate; Color saturation; ELISA; Ultra miniaturized assay