Immunohistochemical analysis of paraffin-embedded human seminoma using Nanog (1E6C4) Mouse mAb #4893 (left) or Mouse (G3A1) mAb IgG1 Isotype Control (right).
Confocal immunofluorescent analysis of normal rat cerebellum using Neurofilament H (RMdO 20) Mouse mAb #2836 (green, left) compared to concentration matched Mouse (G3A1) mAb IgG1 Isotype Control (green, right). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Confocal immunofluorescent analysis of HT-29 cells using α-Tubulin (DM1A) Mouse mAb #3873 (green, left) compared to concentration matched Mouse (G3A1) mAb IgG1 Isotype Control (green, right). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Flow cytometric analysis of MCF7 cells using Pan-Keratin (C11) Mouse mAb #4545 (blue) compared to concentration-matched Mouse (G3A1) mAb IgG1 Isotype Control (red).
Flow cytometric analysis of Jurkat cells, U0126-treated (blue) or TPA-treated (green), using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (E10) Mouse mAb #9106 compared to concentration-matched Mouse (G3A1) mAb IgG1 Isotype Control (red).
Chromatin immunoprecipitations were performed using digested chromatin from HeLa cells and the indicated antibodies. Purified DNA was analyzed by quantitative real-time PCR, using SimpleChIP® Human GAPDH Exon 1 Primers #5516, SimpleChIP® Human RPL30 Exon 3 Primers #7014, and SimpleChIP® Human MYT-1 Exon 1 Primers #4493. The relative abundance of each DNA sequence enriched by Mouse (G3A1) mAb IgG1 Isotype Control (red) is compared to the amount of the same DNA sequence enriched by the histone H3-specific immunoprecipitations (blue).
|Isotype||Mouse IgG1, kappa|
Important! Dilute this control antibody to the same concentration (not dilution) as the specific antibody used for analysis. Higher background may result if excessive amounts of mouse IgG1 isotype control is used.
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.
This protocol is intended for immunoprecipitation of native proteins utilizing Protein G agarose beads for subsequent analysis by western immunoblot or kinase activity.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
10X Cell Lysis Buffer: (#9803) To prepare 10 ml of 1X cell lysis buffer, add 1 ml cell lysis buffer to 9 ml dH2O, mix.
NOTE: Add 1 mM PMSF (#8553) immediately prior to use.
Proceed to one of the following specific set of steps.
NOTE: To minimize masking caused by denatured IgG heavy chains (~50 kDa), we recommend using Mouse Anti-Rabbit IgG (Light-Chain Specific) (L57A3) mAb (#3677) or Mouse Anti-Rabbit IgG (Conformation Specific) (L27A9) mAb (#3678) (or HRP conjugate #5127). To minimize masking caused by denatured IgG light chains (~25 kDa), we recommend using Mouse Anti-Rabbit IgG (Conformation Specific) (L27A9) mAb (#3678) (or HRP conjugate #5127).
posted October 2016
revised April 2018
Protocol Id: 1244
Mouse (G3A1) mAb IgG1 Isotype Control is not directed against any known antigen. It functions as an isotype control for mouse IgG1 monoclonal antibodies.
Isotype control antibodies are used to estimate the nonspecific binding of target primary antibodies due to Fc receptor binding or other protein-protein interactions. An isotype control antibody should have the same immunoglobulin type and be used at the same concentration as the test antibody.
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