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Alkaline phosphatase labeled goat anti rabbit igg

Manufactured by Merck Group
Sourced in United States, China

Alkaline-phosphatase-labeled goat anti-rabbit IgG is a secondary antibody used in immunoassays. It is composed of goat-derived antibodies that specifically recognize and bind to rabbit immunoglobulin G (IgG) molecules. The antibodies are labeled with the enzyme alkaline phosphatase, which can be used to detect and quantify target analytes in various immunoassay applications.

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3 protocols using alkaline phosphatase labeled goat anti rabbit igg

1

Autoantibody Detection in Rabbit Serum

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ELISAs were performed using 4 different neuronal antigens to detect autoantibodies in rabbit serum. Ninety-six well plates were coated overnight in 0.015 M carbonate/0.03 M bicarbonate (pH 9.6) buffer with each of the 4 antigens (Lysoganglioside, tubulin, human dopamine D1 receptor [D1R] antigen, and human dopamine D2 receptor [D2R]) as described previously [48 (link)]. ELISAs were then performed similar to what was described [49 (link)]. In short, washes were performed with PBS containing 0.05% Tween (PBS-Tween) five times and then blocked for 1 h. The wells were washed and sera from immunized or placebo rabbits were diluted and incubated overnight. After five washes, the samples were incubated with alkaline-phosphatase-labeled goat anti-rabbit IgG (Sigma-Aldrich, St. Louis, MO, USA). Then substrate and p-nitrophenyl phosphate were added. The optical density (OD) was measured at 405 nm. The results are expressed as the mean of triplicate wells. All ELISAs were validated with known positive and negative serum controls for each assay to maintain a standardized assay.
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2

AQP4/9 Protein Expression in Spinal Cord Injury

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At 72 hours after spinal cord injury, five rats in each group were sacrificed under anesthesia and spinal cord tissue was harvested. Samples were centrifuged at 1,500 r/min for 30 minutes, and total protein concentration in the supernatant was determined. The samples were then electrophoresed in 5% stacking gel at 40 V for 1 hour and 10% separating gel at 60 V for 3.5 hours, and transferred onto a PVDF membrane at 14 V for 14 hours. The membrane was blocked at 37°C for 2 hours, rinsed three times in TBS for 10 minutes each time, and incubated with rabbit anti-AQP9/AQP4 polyclonal antibody (1:500; Sigma, St. Louis, MO, USA) and rabbit anti-β-actin polyclonal antibody (1:500; Sigma) at room temperature for 60 minutes. After three washes in TBST, samples were incubated with alkaline phosphatase-labeled goat anti-rabbit IgG (1:2,000; Sigma) at room temperature for 60 minutes, developed with DAB (Beijing CellChip Biotechnology Co., Ltd., Beijing, China), and analyzed using Quantity One imaging software (Bio-Rad, Hercules, CA, USA). The AQP4/9 protein expression level was calculated as the integrated optical density ratio of AQP4/9 to β-actin.
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3

Mannan-Induced Complement Activation Assay

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A functional lectin pathway activity assay was done using EDTA-plasma diluted 2.5% in barbital-buffered saline (BBS) incubated for 15′ at 37° C on 10 µg/mL mannan-coated plates.27 (link) C3 deposition was revealed using a polyclonal anti-human-C3c antibody (2.4 µg/mL, Dako) and an alkaline-phosphatase labeled goat anti-rabbit IgG (1 µg/mL, Sigma). Absorption at optical density (OD) 405 nm measured using Infinite M200 spectrofluorimeter (Tecan, CH).
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