Co-Immunoprecipitation of CDC42 and p110β

Co-IP assays were performed as previously described [49 (link)]. Whole-cell protein (500 μg) from organoids was used for co-IP. After overnight incubation at 4 °C with 2 μg antibody, Dynabeads Protein A (Thermo Fisher Scientific, 10002D) was incubated 3 h at 4 °C and then were washed with IP washing buffer three times. Immunoprecipitated proteins were separated by SDS-PAGE and analyzed by Western blot, using antibodies against CDC42 and p110β. The control immunoprecipitation was performed by incubating the lysates with rabbit IgG (Abmart, B30011).

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Jiang R., Tang X., Pan J., Li G., Yang N., Tang Y., Bi S., Cai H., Chen Q., Chen D., Wang H, & Kong S. (2022). CDC42 governs normal oviduct multiciliogenesis through activating AKT to ensure timely embryo transport. Cell Death & Disease, 13(9), 757.

Publication 2022

Dynabeads Protein A B30011

Antibodies Antibody Assays Buffer Cdc42 Cell Immunoprecipitation Organoids P110β Protein a Proteins Rabbit Sds page Western blot

Corresponding Organization :

Other organizations : First Affiliated Hospital of Xiamen University, Third Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University

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Variable analysis

independent variables

Antibody used for co-IP (2 μg)

dependent variables

Co-immunoprecipitation of CDC42 and p110β

control variables

Whole-cell protein (500 μg) from organoids used for co-IP
Overnight incubation at 4 °C
Incubation of Dynabeads Protein A for 3 h at 4 °C
Washing of immunoprecipitated proteins with IP washing buffer three times
Separation of immunoprecipitated proteins by SDS-PAGE
Analysis of immunoprecipitated proteins by Western blot

positive control

Incubation of lysates with rabbit IgG (Abmart, B30011)

negative control

Not explicitly mentioned

Annotations

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