2 replies to this topic

[dcch]

Hello. Just curious to know...In a 80% confluence 24-well plate, what is the normal range for HeLa cell total protein?

Recently I started to harvest cells in 24-well plate for total protein quantification. I typsinized the cells, spin down, wash, then finally added in 50 uL RIPA buffer and shake in cold for 30 min. Then i measured the concentration using Bradford method, and the reading of biophotometer indicated 0.5mg/mL to 1.0mg/mL. I wonder if this protein concentration is normal...

I know the total protein is vary from cells to cells...anyway, can somebody suggest me what should be the "normal range" for the cells harvested from 24-well plate(80%-100% confluency)? So that I can interprete whether my results are acceptable or not...

TQ~

Edited by dcch, 23 September 2009 - 04:02 PM.

[cotchy]

i get around 6-8 mg in total for a confluent T75 of the cells i work with (not HeLa cells)

i am happy with this and think it is about average, i also use ripa etc.

[Dr Teeth]

dcch, on Sep 23 2009, 08:01 PM, said:

Hello. Just curious to know...In a 80% confluence 24-well plate, what is the normal range for HeLa cell total protein?

Recently I started to harvest cells in 24-well plate for total protein quantification. I typsinized the cells, spin down, wash, then finally added in 50 uL RIPA buffer and shake in cold for 30 min. Then i measured the concentration using Bradford method, and the reading of biophotometer indicated 0.5mg/mL to 1.0mg/mL. I wonder if this protein concentration is normal...

I know the total protein is vary from cells to cells...anyway, can somebody suggest me what should be the "normal range" for the cells harvested from 24-well plate(80%-100% confluency)? So that I can interprete whether my results are acceptable or not...

TQ~

Based on Cotchy's response, you should be getting around 0.128-0.170 mg/ml for an 80% confluent 24 well plate. If he obtains only 6-8 mg of protein from a confluent T-75 flask (surface area 7500 mm2), while you are using a 24 well plate (surface area 200 mm2 per well--at 80% confluency, assume surface area of 160 mm2), then you should be getting 6/(7500/160) = 0.128 mg or 8/(7500/160) = 0.170 mg, at least based on the area conversion.
Note: I see now that his values were not from HeLa, so who knows?

Edited by Dr Teeth, 24 September 2009 - 08:45 AM.