Is there a difference between 1% NP-40 versus 1% Nonidet-P40 for RIPA buffer for Co-IP? These are two different detergents but it is a common misconception that they are the same thing. Would one or the other make a difference when trying to run a Co-IP?
I use NP-40 (Pierce Biotech) for lysis of epithelial cells--not Nonidet-P40. The structure of the two is different in the hydrophobic tail, though they have the same polar head group. See this resource for choosing a detergent for RIPA buffer: http://www.piercenet.com/method/detergents-cell-lysis-protein-extraction. I usually see Triton-X-100 and/or Tween20 for RIPA buffers, but your choice of detergent will depend largely on your application and the protein(s) you are interested in studying.
http://en.wikipedia.org/wiki/Nonidet_P-40
Nonidet P-40 is sometimes abbreviated as NP-40, but should not be confused with a different detergent by the same name (NP-40, nonylphenoxypolyethoxyethanol).
I use NP-40 (Pierce Biotech) for lysis of epithelial cells--not Nonidet-P40. The structure of the two is different in the hydrophobic tail, though they have the same polar head group. See this resource for choosing a detergent for RIPA buffer: http://www.piercenet.com/method/detergents-cell-lysis-protein-extraction. I usually see Triton-X-100 and/or Tween20 for RIPA buffers, but your choice of detergent will depend largely on your application and the protein(s) you are interested in studying.
Nonident P40 sold by Sigma is Octylphenoxy)polyethoxyethanol, IGEPAL® CA-630, Octylphenyl-polyethylene glycol where the n value for the polymeric ethylene oxide isnt clearly stated.This detergent is often confused with neutral detergent NP 40 ( not easily commercially available anymore) but also called Nonidet P-40 (NP-40, Shell Chemical Co.) originated from Pierce 136 Biotechnology, Rockford, Il. See Ref: http://chemicalreagent21.wordpress.com/2012/06/04/what-is-nonidet-p-40/
Triton x-100 is a good substitute for this and the n value for the ethylene glycol moeity should be between 9-10.
To confuse matters further; other companies also sell a NP-40 substitute, which may differ in the amount of ethylene glycol as well.
Id suggest you just buy one product from one source and dont change your NP40 between experiments.
Dear Charles,
As Abigail and Aneesa said the detergent use is your choice. I allways tell my students RIPA you compose like pizza... In one recipy you buffer with Tris in other with phosphate or Hepes. Salt concentration you adjust to the ionic strenght your protein likes. With Co-IP you usually have the problem of unspecyfic binding, thus in case of such a problems I suggest to increase salt and detergents concentrations.
best - jan
If you are still looking for your problem please let me know. I can help you in resolving your problem.
There is a 2017 publication on the NP-40 potency subject which has useful information on the history of use of the original Shell NP-40. The difference between the original compound and the substitutes seems to be an extra carbon atom in the hydrophobic tail. The aurhors report that all available NP-40 substitutes were 10x more potent than the 'old school' NP-40.
Use of substitute Nonidet P-40 nonionic detergents in intracellular tubulin polymerization assays for screening of microtubule targeting agents
S. Sinha, J.J. Field, and J.H. Miller
RIPA always causes altered protein profile and endogenous baseline due to loss of proteins non-proportionally. Strong lysis buffer should be used for complete lysis. The stickiness can be removed by using a spin column with a special filter. Sample can be ready in just a minute. Try to search Minute Protein extraction kit on Google.
- Badges
- Science method