Ten µg/ml of Fab-PP form of 1-69Ab was coated onto 96 well Maxisorp immunoplates (Nunc), and K1-18 Ab was added to each well. After incubation with peroxidase-conjugated goat anti-mouse IgG (H+L chain, MBL), HRP substrate (OPD; Wako) was added to each well. After stopping the peroxidase reaction by adding H2SO4, the absorbance of the sample at 492 nm was measured.
Hrp substrate opd
Manufactured by Fujifilm
HRP substrate (OPD) is a chromogenic substrate used in enzyme-linked immunosorbent assays (ELISA) for the detection and quantification of target analytes. The substrate is designed to produce a colored end-product when catalyzed by the horseradish peroxidase (HRP) enzyme, which is commonly conjugated to detection antibodies in ELISA protocols.
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Lab products found in correlation
4 protocols using hrp substrate opd
1
Quantitative Fab-PP Immunoassay
2
ELISA for Formalin-Treated Virus
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Formalin-treated virus particles were coated onto 96 well Maxisorp immunoplates (Nunc), and Fab-cp3 Ab in the supernatant of E. coli culture was added to each well. After incubation with rabbit anti-cp3 Ab (MBL), the wells were further incubated with peroxidase-conjugated goat anti-rabbit IgG (H+L chain; MBL). Then, HRP substrate (OPD; Wako) was added to each well, and the color of the sample was developed. After stopping the peroxidase reaction by adding H2SO4, the absorbance of the sample at 492 nm was measured.
3
Virus Particle Binding Assay
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Formalin-inactivated virus particles (Bri07) were coated onto a 96 well Maxisorp immunoplate. Fab-cp3 molecule that was diluted from 20-hold concentrated solution at an optimized concentration was mixed with an equal volume of 200 or 400 µg/ml of K1-18 Ab and incubated at 37°C for 1 h. The mixture was added to virus coated well and the wells were incubated at 37°C for 1 h. After incubation with rabbit anti-cp3 Ab (MBL), the wells were further incubated with peroxidase-conjugated goat anti-rabbit IgG (H+L chain; MBL). Then, HRP substrate (OPD; Wako) was added to each well, and the color of the sample was developed. After stopping the peroxidase reaction by adding H2SO4, the absorbance of the sample at 492 nm was measured.
4
Competition ELISA for Virus Particle Binding
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Competition ELISA was performed by using the Fab-PP form of Ab [13] (link) or C179 [3] (link) for detection of the binding activity to virus particles and the Fab-cp3 form of Ab as a competitor. Fab-cp3 molecules in the supernatant of E. coli culture were concentrated 20-fold before use. Formalin-inactivated virus particles were coated onto a 96 well Maxisorp immunoplate. A total of 50 µl of Fab-PP at an optimized concentration was mixed with 50 µl of 20-fold concentrated Fab-cp3 and the mixture was added to a virus-coated well. Then, peroxidase-conjugated rabbit anti-streptavidin Ab was added to each well as a secondary Ab. When C179 at the final concentration of 0.25 µg/ml was used for detection of the binding activity to virus strain, each well was incubated with peroxidase-conjugated goat anti-mouse IgG (H+L chain; MBL) as a secondary Ab. Then, HRP substrate (OPD; Wako) was added to each well, and the color of the sample was developed. After stopping the peroxidase reaction by adding H2SO4, the absorbance at 492 nm was measured.
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