triton x and np40 for nuclei isolation
Posted 02 March 2005 - 10:24 PM
I'm currently in the beginning stages of nuclear run on preparations.
For the isolation of the cell nuclei (MCF-7 and MDA-MB-231), the recipe for the lysis buffer is 10mM Tris pH 7.5-8.0, 0.5% NP-40, 10mM NaCl, 3mM MgCl2.
Problem is, we don't have NP-40.... we do have triton X.
I've been googling a bit, but, still would like to have someones opinion on whether these two are interchangable.
Really don't want to screw things up, anymore than what is destined to happen anyway.
Can I use triton X? How much?
Thanks a million in advance.
Posted 03 March 2005 - 02:31 AM
According to the Sigma product info sheet:
In my experience, used at the same concentrations, NP-40 (or Igepal), is more stringent tant TX100. So if you have no option but to use TX100 I would tell you to use 2-3 times the amount of NP40 in your protocol... But I strongly recommend you get some Igepal.
Triton X-100tm has a structure very similar to those of Igepal CA-630 and of Nonidet P-40 (no longer commercially available), and the names are sometimes reported as synonyms. However, Triton X-100 is slightly more hydrophilic than Igepal CA-630; these two detergents are NOT considered to be functionally interchangeable in most applications.
Posted 07 March 2005 - 08:20 AM
I have in the past used Triton-X100 in exchange for NP-40 with no differences in results.
I used to do cell permeabilisations for confocal studies and WB and found that in the presence of 0.3%-0.5% Triton-X100 the nuclear membrane would remain intact however the cytosolic membrane would be permeabilised.
Consequently, by lysing the cytosolic membrane, spinning and then using a nuclear solubilisation buffer usually high salt or the addition of CHAPS or SDS then you could obtain clean nuclear lysates.
Hope this could be of some help
Posted 07 March 2005 - 04:27 PM
Ethylphenolpoly(ethyleneglycolether)x or polyoxyethylene(10)isooctylphenyl ether or t-Octylphenoxypolyethoxyethanol or polyethylene glycol tert-octylphenyl ether (x is approximately 9.5 according to the Sigma catalog) is a non-ionic detergent. polyethylene glycol P-1,1,3,3-tetramethylbutylphenyl ether, octyl phenol ethoxylate, 4-octylphenol polyethoxylate, Mono 30
Triton X-100 is an estrogenic surfactant spermatocide, closely related to Nonoxynol-9. Octylphenol exhibits estrogenic effects in some animals at 30 ppb which is roughly 0.000009% Triton X-100 or one drop in 275 liters of water.
D = 1.070 4-(C8H17)C6H4(OCH2CH2)nOH CAS 9002-93-1
Strictly speaking, Tergitol NP-40 (Union Carbide) previously available as a solid at room temperature as Sigma catalog number NP-40 is not the same as Nonidet-P40 which is a liquid at room temperature ("Nonidet" is a registered trademark owned by Shell Chemicals).
Tergitol type NP-40, (a solid at room temp.) and Nonidet P-40 (no longer available but Sigma sells its equivalent, Igepal CA-630, a liquid at RT).
A recent Fluka catalogue lists 'Nonidet P-40 substitute' with synonyms being NP-40 (as unfortunately Nonidet P-40 was sometimes called) as well as Imbentin-N/52.
Sigma Cat # I3021 IGEPAL CA-630 (octylphenoxy)polyethoxyethanol
“chemically indistinguishable from Nonidet P-40, which is no longer commercially available”