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Huvec cell number ? - (Jul/21/2005 )

I need some help on starting a Western blot.

Can anybody tell me what is the number of cells required to be able to detect your protein of interest when doing a Western blot or Immunoprecipitation ?

I work with cell cultures, specifically Huvec (human umbilical vein endothelial cells).

I am trying to detect for a protein by doing a Western, but don't know what number of cells to grow up to do it.

What would be enough cells ?

ex) grown in 24 or 12 or 6-well plate ?
or in a T-25 or T-75 or T150 flask ?

And what would be a good number of cells seeded in a specific plate ?

Huvec cells don't seem to proliferate very well and takes time to grow to sufficient number.

I would very much appreciate any input.


i use a 6well plate at 80% confluency to do a western.
but the proteins i study are good expressed in the cells.

For IP i use a 10cm plate.