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Western blot analysis of IgM was performed by loading 1ug of purified Human IgM, Human IgA, and Human IgG and 10ul of PageRuler Prestained Protein Ladder (Product # 26616) per well onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane using the G2 Fast Blotter (Product # 62288), and blocked with 5% BSA in TBST for at least 1 hour at room temperature. IgM heavy chain was detected using a mouse anti-human IgM monoclonal antibody (Product # MII0402) diluted to a concentration of 2ug/ml in blocking buffer, overnight at 4C on a rocking platform, followed by an HRP-conjugated goat anti-mouse light chain secondary antibody diluted 1:40,000 for at least 30 minutes at room temperature (panel A). To verify the presence of all isotypes, a second blot was probed with a goat anti-human IgM + IgG +IgA polyclonal antibody (Product # 31128) at a concentration of 0.5ug/ml in blocking buffer overnight at 4C on a rocking platform, followed by an HRP-conjugated mouse anti-goat IgG secondary antibody (Product # 31400) diluted 1:40,000 in blocking buffer for at least 30 minutes at room temperature (panel B). Chemiluminescent detection was performed using SuperSignal West Pico.
|Tested species reactivity||Goat|
|Published species reactivity||Not Applicable|
|Host / Isotype||Mouse / IgG|
|Purification||Antigen affinity chromatography|
|Storage buffer||PBS, pH 7.6, with 15mg/ml BSA|
|Storage Conditions||4° C|
|Cross Adsorption||Against mouse, human and rabbit serum proteins|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||1:500 - 1:5,000|
|Immunocytochemistry (ICC)||1:5,000 - 1:100,000|
|Immunofluorescence (IF)||1:5,000 - 1:100,000|
|Immunohistochemistry (IHC)||1:5,000 - 1:100,000|
|Immunoprecipitation (IP)||1:500 - 1:5,000|
|Western Blot (WB)||1:5,000 - 1:200,000|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
|Western Blot (WB)||See 1 publications below|
Concentration may vary slightly from lot-to-lot, see lot-specific datasheet for exact concentration.
This antibody has been successfully used in Western blot, IF, ICC, IHC, IP and FACS applications.
Antibody Specificity: This antibody reacts with the heavy chains of goat IgG and with light chains common to most goat immunoglobulins. The antibody exhibits minimal inherent cross-reactivity to mouse serum proteins. No antibody was detected against non-immunoglobulin serum proteins. The antibody has been tested by ELISA and/or solid-phase adsorbed to ensure minimal cross-reaction with mouse, human and rabbit serum proteins. However, this antibody may cross-react with immunoglobulins from other species.
Restoration and Storage: Store product at 2-8 °C until opened. Restore with 1.0 ml distilled water (0.8 mg/ml after restoration). Centrifuge product if it is not completely clear after standing for 1-2 hours at room temperature. To judge clarity, draw product into a pasteur pipette. Product may be stored for several weeks at 4°C as an undiluted liquid. After dilution, do not use for more than one day.
To extend the shelf-life of this product, add an equal volume of glycerol to make a final concentration of approximately 50% glycerol and store at -20°C.
Country of Origin: USA
Thermo Scientific Anti-Goat secondary antibodies are affinity-purified antibodies with well-characterized specificity for goat immunoglobulins and are useful in the detection, sorting or purification of its specified target. Secondary antibodies offer increased versatility enabling users to use many detection systems (e.g. HRP, AP, fluorescence). They can also provide greater sensitivity through signal amplification as multiple secondary antibodies can bind to a single primary antibody. Most commonly, secondary antibodies are generated by immunizing the host animal with a pooled population of immunoglobulins from the target species and can be further purified and modified (i.e. immunoaffinity chromatography, antibody fragmentation, label conjugation, etc.) to generate highly specific reagents.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
Poly (ADP-ribose) polymerase inhibitor LT-626: Sensitivity correlates with MRE11 mutations and synergizes with platinums and irinotecan in colorectal cancer cells.
31400 was used in western blot to study the efficacy of a PARP inhibitor in colorectal cancer and the effects of microsatellite instability, MRE11 mutation and co-treatment with platinum compounds or irinotecan
|McPherson LA,Shen Y,Ford JM||Cancer letters (343:217)||2014|