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Pippuri
Member since: Jul 12, 2009
From: Georgia, United States
Status: Protein Chemistry & Glycobiology Moderator
My points: 1053    what's this
Name: [Privacy]
 


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Apr 12, 2012    06:35 AM

Replied to topic in Biochemistry forum   Mechanism of lowering of blood glucose

Hi Sambos, You can find assays for all your enzyme of interested in the following link- Enzyme Explorer/Sigma-Aldrich Good luck.

Mar 29, 2012    09:24 AM

Replied to topic in Protein Detection (Western blots, gels, IP) forum   Western blot mystery - possibly a sample preparation problem?

Hi Mark, I have not used the method you mentioned to prepare sample for WB, so it is pretty hard to tell how it might go wrong. However, I routinely use RIPA buffer to lyse cells and an electronic homoginizer to aid protein extraction from tissue samples as depicted in this link (from Abcam). Al ...

Mar 19, 2012    02:15 PM

Replied to topic in Protein Detection (Western blots, gels, IP) forum   western blotting and immunohistochemistry

Hi sitara, I will say it depends, but for the safe side, (and if you can access to the samples fairly easy), it is better to have both. However, it the antibody and the model that you are working on are both well established, then, western isn't that necessary... Good luck.

Mar 15, 2012    12:26 PM

Replied to topic in Protein Detection (Western blots, gels, IP) forum   trouble in purification with IP

Hi fooooooooox, When using crude material for activity assay, have you already ruled out the possibility of the enzyme activity might come from the endogenous instead of ectopically expressed form? I think that is the first thing you should look into before you start to change tag etc. If you ...

Mar 15, 2012    09:03 AM

Replied to topic in Protein Detection (Western blots, gels, IP) forum   trouble in purification with IP

Hi fooooooooox, Thanks for providing the details. The one possibility is that your enzyme is super-sensitive to acidic condition. How long did you incubate with glycine before the neutralization? May be you have to look into that. Also, can you try to assay for its activity on beads (thus omittin ...

Mar 15, 2012    05:39 AM

Replied to topic in Protein Detection (Western blots, gels, IP) forum   trouble in purification with IP

Hi fooooooooox, Can you be describe your IP conditions (lysis/binding buffer, wash buffer, and elution buffer) in details? Thanks.

Mar 9, 2012    11:18 AM

Replied to topic in Virology forum   help me for selection of best lentiviriseconcentration kit

Hi masomeh_es.haghi, Somebody else in my lab uses it. I don't usually concentrate my lentiviral particles. I found that using "spinoculation" (forcing the pseudo viral particles attached to the cell surface hence enforcing their entry via centrifugation) actually works decently in m ...

Mar 9, 2012    10:43 AM

Replied to topic in Virology forum   help me for selection of best lentiviriseconcentration kit

Hi masoumeh_es.haghi, Here is one without the ultracentrifugation step: Lenti-X concentrator (Clontech) Good luck.

Feb 26, 2012    06:04 AM

Replied to topic in Analytical Chemistry forum   mass spectrometry

Hi ghazaleh842000, There is no reason for one to see singals in MALDI but not in LC-MS/MS. If the peptides are there, they should be able to be detected regardless. Cheers.

Feb 15, 2012    01:38 PM

Replied to topic in Analytical Chemistry forum   mass spectrometry

Hi ghazaleh842000, If the mass spectrometer on which one acquires data has the resolution power, yer, on can tell the charge state of a specific peak- by looking at the distribution of the isotopic variants of that peak (But you really have to zoom-in in order to see that, hence on the first gimp ...

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