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On day 49, the blood was collected from the heart bleeding and centrifuged at 5,000g, and the serum was collected, mixed with 20 folds in volume of KLH solution to preabsorb antiKLH in the antiserum.
For the detection of actin, the membrane was blocked with 1% BSA for 30 minutes at room temperature and probed with the primary antibody for actin (sc-1616, Santa Cruz Biotechnology) (diluted at 1: 500) for overnight at 4[degrees]C and was thereafter incubated with anti-goat IgG, HRP preabsorbed from donkey (ab97120) (1: 5000) for 1 hour at room temperature.
The test was performed as follows: sections of the stomach were incubated with a "working" dilution of the primary immunoserum, which had been previously preabsorbed for 18 h at 37[degrees]C with 20 [micro]g of appropriate purified protein SP (ab120170, Abcam), CGRP (ab158017, Abcam), and L-ENK (ab159087, Abcam).
Then 100 [micro]L/well of preabsorbed serum sample at a 1: 100 dilution in diluting buffer (PBST) was then added and incubated at room temperature for two hours, followed by an additional washing step.
Positive controls for labeling specificity of each peptide were performed by incubating sections with antiserum preabsorbed with the respective antigen (10-100 g/ml).
(12,18,20) Briefly, each threshold optical density was determined by normalizing the data to those of the (preabsorbed negative) control sections.
One is a reaction between the molecules of the target gas and preabsorbed oxygen [4].
Briefly, urine samples (1:20 dilution) or 8-oxo-dGuo standards were first added to an 8-oxo-dGuo/BSA conjugate preabsorbed enzyme immunoassay plate.
In general, there are two types of reactions occurring simultaneously on the oxide surface: [C] the target gas molecules react with the preabsorbed oxygen, and [C] the target gas molecules react directly with the surface atoms of oxide.