Activation of human integrin αIIbβ3 was assessed using the activation-specific human αIIbβ3 antibody PAC1 (BD Bioscience; #340535) and goat anti-mouse IgG secondary antibody Alexa 633 conjugate (Life Technologies; #A-21050). αIIbβ3-CHO cells were developed and cultured as described previously (23 (link)). EGFP-fused talin-H and/or FLAG-tagged DOK-1 expression constructs were transfected into αIIbβ3-CHO cells using Lipofectamine 2000 (Invitrogen, Carlsbad, CA). pEGFP vector (Clontech Laboratories, Mountain View, CA) or pCMV-tag 2B (Stratagene) was transfected as empty vector to adjust the amount of plasmid DNA to be transfected. 24 hours after transfection, the cells were collected and PAC1 binding was analyzed by flow cytometry (FACS LSR II, Becton Dickinson), gating only on the EGFP positive cells.
Protocol detail
Integrin αIIbβ3 Activation Assay
Partial Protocol Preview
This section provides a glimpse into the protocol.
The remaining content is hidden due to licensing restrictions, but the full text is available at the following link:
Access Free Full Text.
Anti igg
Antibody
Cells
Cho cells
Flow cytometry
Goat
Human
Lipofectamine 2000
Mouse
Plasmid
Talin
Transfection
Vector
αiibβ3
Corresponding Organization :
Other organizations : University of Iowa, Harvard University Press, Johns Hopkins University
Top 5 similar protocols
Variable analysis
independent variables
- Transfection of EGFP-fused talin-H and/or FLAG-tagged DOK-1 expression constructs into αIIbβ3-CHO cells
- Activation of human integrin αIIbβ3 assessed using the activation-specific human αIIbβ3 antibody PAC1
- αIIbβ3-CHO cells developed and cultured as described previously
- PEGFP vector or pCMV-tag 2B transfected as empty vector to adjust the amount of plasmid DNA to be transfected
- Positive control: Not explicitly mentioned
- Negative control: Transfection of pEGFP vector or pCMV-tag 2B as empty vector
Annotations
Based on most similar protocols
Etiam vel ipsum. Morbi facilisis vestibulum nisl. Praesent cursus laoreet felis. Integer adipiscing pretium orci. Nulla facilisi. Quisque posuere bibendum purus. Nulla quam mauris, cursus eget, convallis ac, molestie non, enim. Aliquam congue. Quisque sagittis nonummy sapien. Proin molestie sem vitae urna. Maecenas lorem.
Sign up for free or login to display all annotations
As authors may omit details in methods from publication, our AI will look for missing critical information across the 5 most similar protocols.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!