Dear all, good day!
I have a question regarding the use of NaOH for lysis of mammalian cells. Is it a common practice? Are there any publications which have used NaOH to perform cell lysis? If the intention was not to extract proteins/nucleic acid, but to extract a certain chemical compound for LC-MS/MS is it still possible?
For your kind advice, please.
Thank you.
Probably yes, if the "certain chemical compound" object of your search is not destroyed by NaOH, a strong base. If you are looking for an ester, it will have been destroyed.
Affirmative. Sellick et al (DOI 10.1007/s11306-010-0216-9) actually compared alkaline extraction (KOH) for un-targeted metabolomics in mammalian cells. They showed alkali extraction generated 25% fewer metabolites (than methanol) but did enrich specific classes of metabolites. A word of caution: alkaline extractions, specially strong alkali like NaOH or KOH are likely to induce chemical changes like hydrolysis of the ester bonds as Prof. Carlson has emphasized.
NaOH or KOH...not a problem. Need to keep a close eye on Na+ or K+ adducts though and take those into account when performing a library search for untargeted metabolomics especially. Even with targeted assays, please just check for possible adduct formation on the MS side.
Neutralization with HCl at the appropriate pH or even pushing into acidity would help greatly with mass spec detection. All mass specs detect ions at the end of the day...so that's what you need to form (most of the time) before injecting onto the system.
There's definitely papers out there for referencing. Just need to have a good look Justin. Best of luck!!
Justin,
Just about month ago i was looking an answer for something very similar. I worked on a project where we were interested to see uptake of a drug from nanomicelles in retinal cells. And i was searching for protocols enabling cell lysis and drug extraction. What i finally used was acetonitrile with internal standard and 1 freeze thaw cycle. After sample prep the recovery and matrix effect both were in acceptable range. And we are trying to wrap up the study now. At that time i did not know about using high salt concentration. Even if i would know i am not sure if i would use it due to possible Na and K adduct formation which sometimes are huge problems in Ms analysis.
- Badges
- Science topic
- Similar topics
- Linguistics
- Composition Studies
- Publications