I have ITC Data needing interpretations. Can anyone help interpret such data?

10 November 2014 8,561 13 View

How valid are results from ITC with enzymes having multi-substrates?

My enzyme has 3 substrates. I want to determine Kd of each substrates using isothermal titration calorimetry. However, the first substrate does not "bind" with the enzyme unless the second...

07 August 2014 1,574 4 View

What is the next step after poly-alanine molecular replacement?

After a nightmare of MR trials, I've finally got a resonable match using poly-ala model by first pruning (gedit) then chainsaw. Any recommendations of a good site for the next steps would be...

06 July 2014 7,609 5 View

Does anyone know an assay for an enzyme inhibited by O2?

I am convinced that my enzyme is inhibited by atmospheric oxygen. One supporting evidence is that translation of this protein is only seen in hypoxic conditions. I want to support my hypothesis....

02 March 2014 2,557 4 View

Anyone know how to get a clean sample with SEC?

I have tried multiple buffers/conditions/parameters to get a clean sample from size exclusion chromatography SEC using sephadex 200 AKTA fPLC. Anyone have any suggestion how i get a pure protein...

01 February 2014 4,876 6 View

Does anyone know a robust assay?

I need a robust assay to determine enzymatic activity of my protein. The two small molecules are not UV-Vis unique. The two small chemical molecules are Naphthelene backbone and an isoprene tail....

01 February 2014 6,101 5 View

Can anyone help with 260 nm abs for membrane protein?

I purified extensively my membrane protein, ionic exchange, superdex 200, his-bind, etc. SDSPAGE, Bradford test, Western have all shown pure protein protein. This protein is not known to associate...

01 February 2014 1,304 7 View

Membrane Protein Storage conditions?

What glycerol concentration is appropriate for storage of membrane protein in -80? It seems that 10%-30% is causing aggregation after I store it for 10 days.

31 December 2013 10,009 4 View

Is it common that the expected MW of my membrane protein in SDS-PAGE is lower/higher than what is the theoretical size?

Much thanks.

31 December 2013 9,598 20 View

What is the best way to prevent membrane protein aggregation?

I'm expressing a membrane protein, and it seems to be prone to aggregating during purification. I have played with the pI-pH of buffer. What other option(s) are there?

31 December 2013 6,222 40 View

12% ledeburitic cold work tool steel, etched for revealing retained austenite, any help for clarifying?

Hello everyone. Given microstructure image belongs to a 12% ledeburitic cold work tool steel (annealed and tempered). The sample was etched with picral (100 mL ethanol and 4 g picric acid). I...

02 March 2021 4,119 3 View

New Journal, "Integrative Psychological and Behavioral Science" -- do you not know, and have you not seen, this done before?

Question to you and THEM, the New Journal, "Integrative Psychological and Behavioral Science" -- do you not know, and have you not seen, this done before? There appears to be a core problem for...

02 March 2021 3,024 2 View

Does anyone know HRM channel in a 2 plex QIAGEN Rotor- Gene Q qPCR can be used as a separate channel?

I am going to have 3 different probes in my qPCR work that I am going to do. But I realized that the machine we have in the lab is a Rotor-Gene Q 2plex HRM Platform, saying it has green, yellow,...

01 March 2021 8,544 1 View

Why might my GFP tag be dissociating from my plasmid after transfection?

I have a set of stably transfected cell lines all transfected with plasmids containing GFP tags on the C terminus. During a western blot using anti-GFP antibody, one of my plasmids has dissociated...

01 March 2021 9,310 4 View

Precipitate after adding laemmli buffer to protein solution containing Potassium Acetate?

Hi, I am running a size exclusion chromatography experiment with a buffer containing Potassium Acetate as a salt. I analyse these fractions through SDS-PAGE. After boiling my SEC fractions in...

01 March 2021 2,622 3 View

What is the most suitable server for predicting protein structures in bacteria?

I have used the i-Tasser several times, however it has been unavailable for several days. I tried the swiss-model, but the output was not very pleasant due to the model used. Is there any other...

28 February 2021 4,521 3 View

Do you need the CMV enhancer before the CMV promotor for a proper function of CMV on the gene that follows?

I'm a student and I have to produce a cell line with a knockin for the NRF2 gene with GFP. I have to put a promotor in front of the GFP because the gene will be too far away from the promotor of...

28 February 2021 7,127 2 View

Are protein samples in SDS compatible with ELISA?

I have used an AllPrep DNA/RNA/Protein Mini Kit QIAGEN kit to extract RNA and protein from my samples. At the end of the protein purification, I resuspend my protein in 5% SDS. Will these samples...

28 February 2021 7,370 3 View

Unknown debris sheets observed in cell culture flasks?

Hi, We culture neural progenitor cells in T175 culture flasks coated with PLO-Laminin. We have successfully grown these cells with our protocol for months. Recently, we observed large sheets...

25 February 2021 3,443 4 View

How to replicate the well conditions of a crystal hit with 20%(w/v) isopropanol?

I had a crystal hit in a well with 0.1M HEPEs pH 7.5, 10%(w/v) PEG 4000, and 20%(w/v) isopropanol. Why is isopropanol in w/v here if it's just solvent? How do I recreate this well condition?

25 February 2021 9,761 1 View