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Figure 1. Optimization of in-house quantitative sandwich ELISA for detection of HBsAg and HBeAg. Standard curve depicting the optimal dilution of detection antibody for the detection of (A) HBsAg and (B) HBeAg. Primary antibodies used for antigen capture were mouse monoclonal α-HBsAg (Fitzgerald, 10-H05H) and mouse monoclonal α-HBeAg (Fitzgerald, 10-H10M). Detection was achieved using HRP-conjugated α-HBsAg goat <t>polyclonal</t> antibody (Fitzgerald, 70-HG15S) and α-HBeAg mouse monoclonal antibody (Fitzgerald, 61-H10K). OD450 values were normalized against BSA control wells. The assay was optimized by adjusting antibody concentrations to achieve a strong signal-to-noise ratio while maintaining linearity. Data were normalized by subtracting the optical density reading at 450 nanometers (OD450nm) from bovine serum albumin (BSA) control wells. (C) Summary table of ELISA data for HBeAg and HBsAg. Assay quality was assessed based on linearity of the standard curve R2 and Z′ factor. Data from 3 biological replicates.
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Figure 1. Optimization of in-house quantitative sandwich ELISA for detection of HBsAg and HBeAg. Standard curve depicting the optimal dilution of detection antibody for the detection of (A) HBsAg and (B) HBeAg. Primary antibodies used for antigen capture were mouse monoclonal α-HBsAg (Fitzgerald, 10-H05H) and mouse monoclonal α-HBeAg (Fitzgerald, 10-H10M). Detection was achieved using HRP-conjugated α-HBsAg goat <t>polyclonal</t> antibody (Fitzgerald, 70-HG15S) and α-HBeAg mouse monoclonal antibody (Fitzgerald, 61-H10K). OD450 values were normalized against BSA control wells. The assay was optimized by adjusting antibody concentrations to achieve a strong signal-to-noise ratio while maintaining linearity. Data were normalized by subtracting the optical density reading at 450 nanometers (OD450nm) from bovine serum albumin (BSA) control wells. (C) Summary table of ELISA data for HBeAg and HBsAg. Assay quality was assessed based on linearity of the standard curve R2 and Z′ factor. Data from 3 biological replicates.
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Figure 1. Optimization of in-house quantitative sandwich ELISA for detection of HBsAg and HBeAg. Standard curve depicting the optimal dilution of detection antibody for the detection of (A) HBsAg and (B) HBeAg. Primary antibodies used for antigen capture were mouse monoclonal α-HBsAg (Fitzgerald, 10-H05H) and mouse monoclonal α-HBeAg (Fitzgerald, 10-H10M). Detection was achieved using HRP-conjugated α-HBsAg goat polyclonal antibody (Fitzgerald, 70-HG15S) and α-HBeAg mouse monoclonal antibody (Fitzgerald, 61-H10K). OD450 values were normalized against BSA control wells. The assay was optimized by adjusting antibody concentrations to achieve a strong signal-to-noise ratio while maintaining linearity. Data were normalized by subtracting the optical density reading at 450 nanometers (OD450nm) from bovine serum albumin (BSA) control wells. (C) Summary table of ELISA data for HBeAg and HBsAg. Assay quality was assessed based on linearity of the standard curve R2 and Z′ factor. Data from 3 biological replicates.

Journal: Pathogens (Basel, Switzerland)

Article Title: Development of Low-Cost In-House Assays for Quantitative Detection of HBsAg, HBeAg, and HBV DNA to Enhance Hepatitis B Virus Diagnostics and Antiviral Screening in Resource-Limited Settings.

doi: 10.3390/pathogens14030258

Figure Lengend Snippet: Figure 1. Optimization of in-house quantitative sandwich ELISA for detection of HBsAg and HBeAg. Standard curve depicting the optimal dilution of detection antibody for the detection of (A) HBsAg and (B) HBeAg. Primary antibodies used for antigen capture were mouse monoclonal α-HBsAg (Fitzgerald, 10-H05H) and mouse monoclonal α-HBeAg (Fitzgerald, 10-H10M). Detection was achieved using HRP-conjugated α-HBsAg goat polyclonal antibody (Fitzgerald, 70-HG15S) and α-HBeAg mouse monoclonal antibody (Fitzgerald, 61-H10K). OD450 values were normalized against BSA control wells. The assay was optimized by adjusting antibody concentrations to achieve a strong signal-to-noise ratio while maintaining linearity. Data were normalized by subtracting the optical density reading at 450 nanometers (OD450nm) from bovine serum albumin (BSA) control wells. (C) Summary table of ELISA data for HBeAg and HBsAg. Assay quality was assessed based on linearity of the standard curve R2 and Z′ factor. Data from 3 biological replicates.

Article Snippet: The α-HBsAg goat polyclonal detection antibody used in the sandwich ELISA (70-HG15S, Fitzgerald, Acton, MA, USA) was prepared in advance by conjugating HRP at 4:1 (antibody-to-HRP ratio) using a commercial kit (LNK002P, Bio-Rad, Hercules, CA, USA).

Techniques: Sandwich ELISA, Control, Enzyme-linked Immunosorbent Assay, HBsAg Assay