western blot

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Blur gal
Blur gal's picture
western blot

 Hi. western blot is a new technique for me as my work usually revolves around genomics microarrray. So, my lack of experience makes my work a bit difficult.

1) May I know what type of antibody needs to be used for primary and secondary? My guess is, if my primary antibody is from a mouse, so my secondary antibody should be from mouse source as well.
2) Is it advisable to to re-use the same mebrane for other antibody for up to three times? 
3)If I am to set up a knockdown experiment, I need both protein and RNA. Is there any particular reason that I may need a bigger dish (i.e. P90) for protein and smaller (i.e. P60) dish for RNA?

Thank you.

Blur gal

shankaranarayanan.v
shankaranarayanan.v's picture
Hi blur gal

Hi blur gal

1. in western blots if your primary antibody is derived from mouse, the secondary antibody you should be using is anti-mouse. or the same for any other like primary rabbit, secondary anti - rabbit

2. yes, you can re- use the antibodies more than once..

3. as for as your third question, i don't have much idea abt it.

regards,
Shankar

Blur gal
Blur gal's picture
 hi,

 hi,

thank you for your reply. :)