Dear,
Anyone has/tried some protocol for extraction of whole cell from rat testis for western blot? Which buffer you used, troubleshooting and similar things.
Dear Milos,
Please read the following:
Testis (Mouse) Whole Cell Lysate - normal tissue (ab29289)
Overview
Product nameTestis (Mouse) Whole Cell Lysate - normal tissue
See all Testis lysates
Tested applicationsWBmore details
Properties
FormLiquid
Storage instructionsStore at -80°C. Allow to warm to room temp and agitate gently before aliquotting.
Storage bufferPreservative: None
Constituents: 4% SDS, 10% Glycerol, 0.1% Bromophenol blue, Protease inhibitor, Sodium chloride, EDTA, Magnesium chloride, HEPES, 0.15M Beta mercaptoethanol
Concentration1 mg at 3.75 mg/ml
Lysate notesProtein is isolated from whole tissue homogenates using a proprietary technique. The tissue was frozen in liquid nitrogen immediately after excision and then stored at -70ºC. Quality control: 1. The isolated protein pattern on SDS-PAGE gel is visualized by coomassie blue staining. The pattern is consistent with each lot. 2. The isolated protein is Western Blot analyzed by either a GAPDH or beta-actin antibody. The expression level is consistent with each lot.
Research areasKits/ Lysates/ Other
Lysates
Whole Cell Lysates
Mouse
Genito-Urinary
Applications
Our Abpromise guarantee covers the use of ab29289 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application
Abreviews
Notes
WB
Use at an assay dependent dilution. Samples need to boiled for 5 minutes and put on ice before use.
http://www.abcam.com/testis-mouse-whole-cell-lysate-normal-tissue-ab29289.html
Hoping this will be helpful,
Rafik
Dear Milos,
Please read the following:
Testis (Mouse) Whole Cell Lysate - normal tissue (ab29289)
Overview
Product nameTestis (Mouse) Whole Cell Lysate - normal tissue
See all Testis lysates
Tested applicationsWBmore details
Properties
FormLiquid
Storage instructionsStore at -80°C. Allow to warm to room temp and agitate gently before aliquotting.
Storage bufferPreservative: None
Constituents: 4% SDS, 10% Glycerol, 0.1% Bromophenol blue, Protease inhibitor, Sodium chloride, EDTA, Magnesium chloride, HEPES, 0.15M Beta mercaptoethanol
Concentration1 mg at 3.75 mg/ml
Lysate notesProtein is isolated from whole tissue homogenates using a proprietary technique. The tissue was frozen in liquid nitrogen immediately after excision and then stored at -70ºC. Quality control: 1. The isolated protein pattern on SDS-PAGE gel is visualized by coomassie blue staining. The pattern is consistent with each lot. 2. The isolated protein is Western Blot analyzed by either a GAPDH or beta-actin antibody. The expression level is consistent with each lot.
Research areasKits/ Lysates/ Other
Lysates
Whole Cell Lysates
Mouse
Genito-Urinary
Applications
Our Abpromise guarantee covers the use of ab29289 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application
Abreviews
Notes
WB
Use at an assay dependent dilution. Samples need to boiled for 5 minutes and put on ice before use.
http://www.abcam.com/testis-mouse-whole-cell-lysate-normal-tissue-ab29289.html
Hoping this will be helpful,
Rafik
Dear Rafik,
My intention was not to use commercial kit for isolation and preparation of proteins, but to do it manually. Which is buffer composition with exact molar concentrations?
Sincerely,
Milos
Dear Milos,
My intention was that you look at the buffer suggested by Sigma for their product.
Rafik
Hi Milo,
Our lab does whole testis lysates as well as seminiferous epithelium lysates. You can see our protocol in this paper in the materials and methods under immunoblotting. We generally use RIPA buffer but the seminiferous epithelium isolation requires a bit more work and the technique is referenced in the methods there http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3507196/?tool=pmcentrez
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