We used serial plasma dilutions (10%, 5%, 2.5%, 1.25%, and 0.75% in BBS) and pooled plasma of mice belonging to the same experimental group, incubating the samples for 15 min on mannan- (10 µg/mL) or acBSA-coated (25 µg/mL) plates prepared according to a procedure reported previously.18 (link) Plates were washed and incubated for 1 h 30 min at RT with a polyclonal anti-human-C3c antibody (Dako, A0062) diluted to 2.4 µg/mL in wash buffer. After washing, plates were incubated with an alkaline phosphatase-labeled goat anti-rabbit IgG antibody (Sigma A-3812) diluted to 1 µg/mL in wash buffer for 1 h 30 min at RT. After washing, the assay was developed by adding 100 µL of substrate solution (Sigma Fast p-Nitrophenyl Phosphate Tablets, Sigma). The absorbance at 405 nm was then measured using the Infinite M200 spectrofluorimeter managed by Magellan software (Tecan, CH).
Alkaline phosphatase labeled goat anti rabbit igg antibody
Manufactured by Merck Group
Alkaline phosphatase-labeled goat anti-rabbit IgG antibody is a laboratory reagent used to detect and quantify rabbit immunoglobulin G (IgG) in various analytical techniques. The antibody is produced in goats and is conjugated with the enzyme alkaline phosphatase, which can be used to generate a colorimetric or chemiluminescent signal when exposed to the appropriate substrate.
Automatically generated - may contain errors
Lab products found in correlation
Anti human c3c antibody, by Agilent Technologies (1 mentions)
Infinite m200 spectrofluorimeter, by Tecan (1 mentions)
Sigmafast p nitrophenyl phosphate tablets, by Merck Group (1 mentions)
A0062, by Agilent Technologies (1 mentions)
Fast p nitrophenyl phosphate tablets, by Merck Group (1 mentions)
Infinite m200, by Tecan (1 mentions)
5 bromo 2 deoxyuridine brdu labeling reagent, by Roche (1 mentions)
Anti akt antibody, by Cell Signaling Technology (1 mentions)
Goat anti mouse igg antibody, by Merck Group (1 mentions)
3 protocols using alkaline phosphatase labeled goat anti rabbit igg antibody
1
Quantifying Plasma C3c Complement Binding
2
Quantifying Cell-Associated C3c Levels
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Fixed cells were washed three times with washing buffer (10 mM Tris-HCl, 140 mM NaCl, 5 mM CaCl2, 0.05% Tween 20, pH 7.4) and incubated with BSA (1% in washing buffer) for 2 h at RT. After three washings, cells were incubated with a rabbit polyclonal anti-human C3c (Dako, A0062) diluted 1:5000 in washing buffer, for 1 h 30 min at RT. After washing, cells were incubated with an alkaline-phosphatase-labeled goat anti-rabbit IgG antibody (Sigma A-3812) diluted 1:1000 in washing buffer for 1 h 30 min at RT. Cells were then washed and the assay was developed using substrate solution (Fast p-Nitrophenyl Phosphate tablets, Sigma). The absorption was read at OD 405 nm using the spectrofluorimeter (TECAN plate reader, Infinite M200, Switzerland).
3
Cell Proliferation and Signaling Assays
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Sp was purchased from Sigma-Aldrich (St. Louis, MO, USA). 5-Bromo-2′-deoxyuridine (BrdU) Labeling reagent and the Detection Kit I were obtained from Roche Diagnostics GmbH (Mannheim, Germany). Anti-cyclin D1 and anti-p27 antibodies were from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA). Anti-mitogen-activated protein kinase (MAPK), anti-PI3K p85 and anti-AKT antibodies were all purchased from Cell Signaling Technology, Inc. (Danvers, MA, USA). Alkaline phosphatase-labeled goat anti-rabbit IgG antibody and goat anti-mouse IgG antibody were both from Sigma-Aldrich.
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