Western Blot Analysis for Tissue

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Author:Dang Vu Hoang
Source:Laboratory of Vet.Biochemistry and Molecular Biology, Chungbuk National University, Republic of Korea

Abstract: This protocol was used for tissues in detail to detect signals of protein

Preparation of Sample

1. Protein extraction

-Preparation of Tissue: 10-20 mg

-Homogenize sample in 0.6 ml PRO-PREP solution by using homogenizer.

-Incubate at –20℃ for overnight.

-Centrifuge at 14.000 rpm for 10 mins at 40C and transfer supernatant to a fresh tube (1.5 ml tube).

2. Protein measurement

-Prepare sample solution: 50 ml

2 ml Protein plus 48 ml DW (dilution factor: 2)

-Prepare 1X Pro-Measure solution: Dilute 1 part of 10X Pro-Measure solution by adding 9 parts of DW. Vortex sufficiently.

-Add 500 ml 1X Pro-Measure solution into 50 ml sample solution. Vortex sufficiently.

-Incubate for 2-5 mins at RT.

-Measure absorbance at 595 nm (500 ml).

-Adjust Protein concentration: 6 mg/ml.

-Re-measure.