Cat. # | Size | Qty. | Price |
---|---|---|---|
4465T | 20 µl |
|
|
4465S | 100 µl |
|
REACTIVITY | H |
SENSITIVITY | Endogenous |
MW (kDa) | 52 |
Source/Isotype | Rabbit IgG |
Product Information
Application | Dilution |
---|---|
Western Blotting | 1:1000 |
Immunofluorescence (Immunocytochemistry) | 1:50 - 1:200 |
Flow Cytometry (Fixed/Permeabilized) | 1:100 - 1:400 |
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Loading of prestained molecular weight markers (#59329, 10 µl/lane) to verify electrotransfer and biotinylated protein ladder (#7727, 10 µl/lane) to determine molecular weights are recommended.
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised June 2020
Protocol Id: 10
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.
Recommended Fluorochrome-conjugated Anti-Rabbit secondary antibodies:
NOTE: Cells should be grown, treated, fixed and stained directly in multi-well plates, chamber slides or on coverslips.
NOTE: All subsequent incubations should be carried out at room temperature unless otherwise noted in a humid light-tight box or covered dish/plate to prevent drying and fluorochrome fading.
posted December 2010
Protocol Id: 3
All reagents required for this protocol may be efficiently purchased together in our Intracellular Flow Cytometry Kit (Methanol) #13593, or individually using the catalog numbers listed below.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
NOTE: When including fluorescent cellular dyes in your experiment (including viability dyes, DNA dyes, etc.), please refer to the dye product page for the recommended protocol. Visit www.cellsignal.com for a full listing of cellular dyes validated for use in flow cytometry.
NOTE: Adherent cells or tissue should be dissociated and in single-cell suspension prior to fixation.
NOTE: Optimal centrifugation conditions will vary depending upon cell type and reagent volume. Generally, 150-300g for 1-5 minutes will be sufficient to pellet the cells.
NOTE: If using whole blood, lyse red blood cells and wash by centrifugation prior to fixation.
NOTE: Antibodies targeting CD markers or other extracellular proteins may be added prior to fixation if the epitope is disrupted by formaldehyde and/or methanol. The antibodies will remain bound to the target of interest during the fixation and permeabilization process. However, note that some fluorophores (including PE and APC) are damaged by methanol and thus should not be added prior to permeabilization. Conduct a small-scale experiment if you are unsure.
NOTE: Count cells using a hemocytometer or alternative method.
posted July 2009
revised June 2020
Protocol Id: 404
Human
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues near the carboxy terminus of human keratin 7 protein.
Keratins (cytokeratins) are intermediate filament proteins that are mainly expressed in epithelial cells. Keratin heterodimers composed of an acidic keratin (or type I keratin, keratins K9-K28) and a basic keratin (or type II keratin, keratins K1-K8 and K71-K80) assemble to form filaments. Keratin isoforms demonstrate tissue- and differentiation-specific profiles that make them useful as research and clinical biomarkers (1,2).
Dysregulation/mutations in keratin genes can lead to a variety of disorders affecting the skin, hair, nails, and other epithelial tissues (3). While expression of keratins can be variable, immunohistochemical staining of keratins is widely used to help in the identification and classification of epithelial tumors, and may also provide prognostic information.
Keratins 8 and 18 (K8/K18) are expressed in simple epithelia of normal tissue, as well as in adenocarcinomas of the breast, lung, ovary, and gastrointestinal tract. Keratin 17 is expressed in basal keratinocytes of stratified epithelia, hair follicles, and sebaceous glands. Onset of keratin 17 expression coincides with the definition of major epithelial lineages during skin development (4). Keratin 14 (K14) is expressed in basal cells of stratified epithelia, and in basal-like subtypes of breast cancer and squamous cell carcinomas. Keratin 19 (K19) is expressed in glandular epithelia, including the liver, gallbladder, and pancreas, as well as in adenocarcinomas of the breast, thyroid, and bile duct. Keratin 20 (K20) is expressed in gastrointestinal epithelium, urothelium, and Merkel cells in the skin, as well as in colorectal carcinomas and some urothelial carcinomas. Keratin 5/6 (K5/6) is expressed in basal cells of stratified epithelia, including the skin, prostate, and breast, as well as in basal-like breast cancers, squamous cell carcinomas, and some lung carcinomas. Keratin 7 (K7) is expressed in glandular epithelia, such as those in the lung, breast, and female reproductive tract, as well as in adenocarcinomas of the lung, breast, and ovary (5,6).
Keratins, particularly K8, K18, and K19, serve as biomarkers for identification of circulating tumor cells (CTCs) (5).
Post-translational modifications, including phosphorylation, acetylation, ubiquitylation, sumoylation, glycosylation, and transamidation, have been shown to affect the functions of keratins in normal and disease states (6). Understanding the molecular mechanisms underlying these PTMs may provide insights into cancer pathogenesis.
Keratin 7 and Keratin 19 are present in hepatic and pancreatic progenitor/stem cells (7,8).
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