A Paper-Based Immunoassay for the Detection of Immunoglobulin G

Published: March 29, 2024

Abstract

Source: Peng, Y. et al., Covalent Binding of Antibodies to Cellulose Paper Discs and Their Applications in Naked-eye Colorimetric Immunoassays. J. Vis. Exp. (2016)

The video demonstrates a paper-based immunoassay, involving amine-functionalized cellulose discs cross-linked with glutaraldehyde. Covalent immobilization of IgG-Fc-specific capture antibodies, followed by the introduction of immunoglobulin G and horseradish peroxidase-conjugated detection antibodies, results in a color change confirming the presence of immunoglobulin G.

Protocol

1. Grafting Amine Functional Groups on Cellulose Paper Discs Prepare one piece of square paper with a dimension of 1 cm × 1 cm, and 100 paper discs made from grade No. 1 cellulose paper with a diameter of 6.0 mm (medium-flow filter paper) using a hole punch. To derive -NH2 groups on the paper discs, mix 1 ml 3-aminopropyltrimethoxysilane (APS) and 10 ml acetone in a 50 ml glass bottle in the fume hood. Add paper discs to the freshly prepared APS reagent mixture,…

Representative Results

Figure 1. Fourier transform infrared (FTIR) spectra of untreated and APS-treated medium-flow filter square paper (A) and fast-flow filter square paper (B). A. The spectra for untreated medium-flow filter square paper was similar to that of APS treated medium-flow filter square paper. The increase in intensities at bands of 902-1,170 cm-1&#160…

開示

The authors have nothing to disclose.

Materials

Cellulose filter paper, Grade1 (medium flow filter paper) GE Healthcare Pte Ltd Singapore  1001 110
Cellulose filter paper, Grade113 (Fast flow filter paper) Sigma-Aldrich, Singapore 1113-320
3-Aminopropyltrimethoxysilane Sigma-Aldrich, Singapore T1255 >96%
Glutaraldehyde Sigma-Aldrich, Singapore G6257 Grade II, 25% in H2O
Bovine serum album  Sigma-Aldrich, Singapore A2153
Skimmed milk powder Louis François  Packed by Kitchen Capers, Singapore
Tris base Promega H5135
Sodium periodate Merck 106597
Na2HPO4 Merck 106585
KH2PO4 Merck 104873
NaCl CALBIOCHEM 567441
NaOH Merck 106462
HCL Merck 100317
Phosphate buffer saline (PBS) N/A N/A PBS, containing 137 mmol/L NaCl, 2.7 mmol/L KCl, 8.0 mmol/L Na2HPO4 and 1.5 mmol/L KH2PO4, is prepared with water and adjusted to pH 7.4 with 0.1 mol/L NaOH or 0.1 mol/L HCl
Acetone Tee Hai Chem Pte Ltd Singapore 9005-68
Mixture of TMB and hydrogen peroxide solution  1-Step ultra TMB-ELISA solution , Thermo Scientific Pierce 34029 1 L
Affinity purified goat anti-Mouse IgG-Fc coating antibody Bethyl Laboratories, Inc A90-131A
Mouse reference serum Bethyl Laboratories, Inc RS10-101-5 9.5 mg/mL
HRP conjugated goat anti-mouse IgG-Fc detection antibody Bethyl Laboratories, Inc A90-131P
Equipment
Fourier transform infrared spectrophotometer Shimadzu IR Prestige-21  N/A
Oven NUVE FN500 N/A
Turbo mixer VM-2000 MYC LTD N/A
Image J RGB, free download N/A

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記事を引用
A Paper-Based Immunoassay for the Detection of Immunoglobulin G. J. Vis. Exp. (Pending Publication), e22100, doi: (2024).

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