Goat anti-Mouse IgG2a Cross-Adsorbed, Alexa Fluor™ 647 (A-21241) (2024)

Multiplexed fluorescent western blot was performed using Goat anti-Mouse IgG2a Cross-Adsorbed Secondary Antibody, Alexa Fluor™ 647 (Product # A-21241). Whole cell extracts of U-87 MG (Lane 1, 2, 3), SH-SY5Y (Lane 4, 5), HeLa (Lane 6) and HepG2 (Lane 7) were electrophoresed usingNuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP03222BOX). Resolved proteins were transferred onto anitrocellulose membrane (Produc... View More {{ $ctrl.currentElement.advancedVerification.fullName }} validation info. View more

• Alexa Fluor 350
• Alexa Fluor 488
• Alexa Fluor 546
• Alexa Fluor 555
• FITC
• PE

To minimize cross-reactivity, these goat anti-mouse IgG2a whole secondary antibodies have been affinity purified and cross-adsorbed against mouse IgM, mouse IgA, pooled human sera, purified human paraproteins, and mouse isotypes IgG1, IgG2b, and IgG3 prior to conjugation. Cross-adsorption or pre-adsorption is a purification step to increase specificity of the antibody resulting in higher sensitivity and less background staining. The secondary antibody solution is passed through a column matrix containing immobilized serum proteins from potentially cross-reactive species. Only the nonspecific-binding secondary antibodies are captured in the column, and the highly specific secondaries flow through. The benefits of this extra step are apparent in multiplexing/multicolor-staining experiments (e.g., flow cytometry) where there is potential cross-reactivity with other primary antibodies or in tissue/cell fluorescent staining experiments where there are may be the presence of endogenous immunoglobulins.

Alexa Fluor dyes are among the most trusted fluorescent dyes available today. Invitrogen™ Alexa Fluor 647 dye is a near-infrared-fluorescent dye with excitation ideally suited to the 647 nm laser line. For stable signal generation in imaging and flow cytometry, Alexa Fluor 647 dye is pH-insensitive over a wide molar range. Probes with high fluorescence quantum yield and high photostability allow detection of low-abundance biological structures with great sensitivity. Alexa Fluor 647 dye molecules can be attached to proteins at high molar ratios without significant self-quenching, enabling brighter conjugates and more sensitive detection. The degree of labeling for each conjugate is typically 2-8 fluorophore molecules per IgG molecule; the exact degree of labeling is indicated on the certificate of analysis for each product lot.

Using conjugate solutions: Centrifuge the protein conjugate solution briefly in a microcentrifuge before use; add only the supernatant to the experiment. This step will help eliminate any protein aggregates that may have formed during storage, thereby reducing nonspecific background staining. Because staining protocols vary with application, the appropriate dilution of antibody should be determined empirically. For the fluorophore-labeled antibodies a final concentration of 1-10 µg/mL should be satisfactory for most immunohistochemistry and flow cytometry applications.

Product will be shipped at Room Temperature.