1 × 106 cells were lysed directly into 2.5x Laemmli sample buffer, separated by 8% SDS-PAGE, and transferred to PVDF-FL membranes (Millipore). Membranes were blocked in 5% milk in PBS and incubated with primary antibody diluted in 5% milk in PBS supplemented with 0.1% Tween20. Secondary antibodies were diluted in 5% milk in PBS supplemented with 0.1% Tween20 and 0.01% SDS. Membranes were imaged with a Licor Odyssey instrument. Primary antibodies used in these experiments were rabbit anti-Cas9 (Abcam ab204448) and mouse anti-HSP90 (BD Transduction Laboratories 610418). Secondary antibodies used were goat anti-rabbit IRdye 800CW (Licor 827-08365) and goat anti-mouse Alexa Fluor 680 (Molecular Probes A-21057). Relevant regions of each immunoblot are shown in Figs 1 and 3 , and full images are provided in the supplement.
Ab204448
Manufactured by BD
The Ab204448 is a laboratory equipment product. It is designed to perform a core function related to the research and analysis in a laboratory setting. No further details about the intended use or specific capabilities of this product are available at this time.
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Lab products found in correlation
2 protocols using ab204448
1
Western Blot Analysis of Cas9 Protein
2
Quantitative Western Blot Analysis
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1 × 106 cells were lysed directly into 2.5x Laemmli sample buffer, separated by a 4–20% gradient SDS-PAGE gel, and transferred to PVDF-FL membranes (Millipore). Membranes were blocked in 5% milk in PBS and incubated with primary antibody diluted in 5% milk in PBS supplemented with 0.1% Tween20. Secondary antibodies were diluted in 5% milk in PBS supplemented with 0.1% Tween20 and 0.01% SDS. Membranes were imaged with a LICOR Odyssey instrument. Primary antibodies used in these experiments were rabbit anti-Cas9 (Abcam ab204448) and mouse anti-HSP90 (BD Transduction Laboratories 610418). Secondary antibodies used were goat anti-rabbit IRdye 800CW (LICOR 827-08365) and goat anti-mouse Alexa Fluor 680 (Molecular Probes A-21057).
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