Lung tissues were homogenized in RIPA lysis buffer containing phosphatase inhibitors (Santa Cruz Biotechnology Inc., Santa Cruz, CA, USA). Two milligrams of each lysate were applied to 12.5% sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to Hybond-P membranes (GE Healthcare Ltd, Buckinghamshire, UK). After blocking with Blocking One-P (Nacalai Tesque Inc, Kyoto, Japan), the membranes were incubated with anti-AQP5 (GTX11586, x2000, GeneTex, Inc., Irvine, CA, USA) or anti-β-actin (PM053, 1:1000, MBL Co., Nagoya, Japan). They were washed and incubated with peroxidase-conjugated anti-rabbit IgG (1:5000, MBL Co.). The protein bands were visualized with Chemi-Lumi reagents (Nacalai Tesque Inc) and image-captured with a CCD camera system, ImageQuant LAS 4000 mini (GE Healthcare Ltd).
Chemi lumi reagents
Manufactured by Nacalai Tesque
Sourced in Japan
Chemi-Lumi reagents are a series of chemical solutions used in chemiluminescence-based detection techniques. These reagents are designed to facilitate the measurement of light emissions produced by a chemical reaction, which can be correlated with the presence or quantity of specific target analytes. The core function of Chemi-Lumi reagents is to provide the necessary components for the chemiluminescent process, enabling researchers and laboratories to accurately detect and quantify a variety of biomolecules, enzymatic activities, and other analytes of interest.
Automatically generated - may contain errors
2 protocols using chemi lumi reagents
1
Quantification of Aquaporin-5 in Lung Tissue
2
Western Blot Analysis of Thyroid Proteins
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Thyroid tissues were homogenized in RIPA lysis buffer containing phosphatase inhibitors (Santa Cruz Biotechnology Inc., Santa Cruz, CA, USA). Two μg of each lysate were applied to 10% SDS-PAGE and transfer to Hybond-P membranes (GE Healthcare Ltd, Buckinghamshire, UK). After blocking with skim milk, they were incubated with anti-Tpo (MoAb47, 1:250, Santa Cruz Biotechnology Inc.), anti-TG (2H11 + 6E1, 1:400, Novus Biologicals, Centennial, CO, USA), or anti-GAPDH (3H12, 1:1000, MBL Co., Nagoya, Japan). They were washed and incubated with peroxidaseconjugated anti-mouse IgG (1:5000, MBL Co.). The protein bands were visualized with Chemi-Lumi reagents (Nacalai Tesque Inc., Kyoto, Japan) and image-captured with a CCD camera system, ImageQuant LAS 4000mini (GE Healthcare Ltd). The band intensities were quantified using ImageJ analysis software (http://imagej.nih.gov).
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