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Questions related from Sandeep Kumar
I have performed a single treatment (25 micro gram) of a bacterial virulence factor in presence of multiple inhibitors to see the effect in single cell line. Untreated cells, H2O2, virulence...
14 September 2021 3,643 4 View
I have taken H2DCFDA fluoresce at 485/535 and now I want make a ROS relative production bar graph for values. Kindly suggest different steps.
22 February 2020 8,388 3 View
I'm using transparent polystyrene 96 well plate for H2dcfda assay, is it ok or I should use black plate with transparent base.
01 February 2020 774 4 View
I'm using h2o2 as a positive control for h2dcfda assay for the detection of ROS. There's five fold less (20%) Ros than untreated control (100%) Thp1 cells. I'm treating cells for 24 Hours. Can I...
28 January 2020 7,508 14 View
I'm treating Thp1 cells with 10 ng pma and incubating for 24 hours. Is it ok or I should incubate them for more. Is it ok to give treatment immediate after PMA treatment or we should incubate in...
27 January 2020 2,150 5 View
My m values are coming like 237, 286 or 2276 for y=mx+c during bradford assay, due to which my protein concentration is becoming very low. My R square values are around 0.97 or 0.98 or 0.95 and...
09 November 2019 9,701 3 View
Hello everyone, I'm digesting 2 ug of midiprep isolated pET15b and 22b with BamHI and XhoI at 37 oC for overnight. There is smearing in gel and size of the cut vectors also not appearing actual,...
07 June 2019 4,550 3 View
I'm analyzing 445 protein sequences of a single protein for phylogenetic analysis. Multiple sequence alignment of these sequences is showing gaps of different sizes. Is it ok to keep these gaps...
29 April 2019 1,181 3 View
I'm using 0.7% agarose gel for elution of restriction digested plasmid DNA and my insert ranging from 2.3 kb to 5.7 kb from two different brands having gel strength >1200g/cm2 (1.5% ) and...
24 October 2018 1,244 4 View
I have gel purified double digested vector and insert having concentration 1 ng/ul and 6 ng/ul respectively. I have tried ligation thrice at room temperature but did not get any colonies. I'm...
03 October 2018 3,640 20 View
I have isolated pET15b and pET28a, run on 1% agarose gel, they are showing the size exactly around 5.7 and 5.4 kb. I'm just wondering when I will digest them what will be their size and which one...
31 August 2018 3,454 4 View
I'm transforming CaCl2 competent 50 ul E. coli DH5alpha with 1 ul of pET28a and pET30 at 42 oC for 45 seconds. I added 1 ml LB and incubate at 37 oC with shaking for 1 hour and then plated on LB...
28 August 2018 1,487 5 View
What is the importance of Kozak sequence in protein expression during cloning?
20 July 2018 6,678 4 View
Is there any method through which we can perform MTT assay for different time points in a single 96 well plate. I'm treating cells with six different concentration and now want to see there...
31 May 2018 6,342 4 View
We have an ELISA plate reader in our lab having an option path length correction. I'm just wondering when we should use this function and how it works.
22 May 2018 1,717 3 View
We have 500 ml Q sepharose in 20% ethanol in our lab. which is forming solid mass when we store it 4 oC after using. Is it a common property of this resin or it has been expired. It separates...
19 May 2018 6,835 3 View
I'm diluting 1, 2, 4, 8, 16, 32 & 64 ul of 10mg/ml of BSA solution in to 99, 98, 96, 92, 84, 68 & 36 ul of PBS buffer, making total 100 ul. Now adding 50 ul of Bradford reagent. I'm...
28 January 2018 5,896 3 View
I want to clone a 1290 aa long bacterial secretory protein having signal sequence (33 aa), autotransporter domain (at C terminal) and hydrophobic domain (acts for main function of protein) and...
19 December 2017 907 3 View
I want to isolate a protein through anion exchange chromatography using Q sepharose. The theoretical PI of protein is 9.02, Molecular weight is 139312.18, Number of amino acids is 1290, Total...
05 November 2017 2,998 3 View
We want to purchase two workstations for plants and microbial metagenomics, molecular dimension (MD) simulations and docking studies of macromolecules. What would be the best specifications and...
04 November 2017 5,003 4 View
I need to degas buffers for column chromatography, how we can perform it in a lab without any sophisticated instruments. We have a small vacuum pump.
04 November 2017 4,718 3 View
I want to know surface charge of a protein of 1290 aa long for ion exchange chromatography isolation. Kindly suggest me how to calculate it.
09 October 2017 4,774 4 View
I want to eliminate redundant and incomplete sequences from my PSI-BAST result of a protein for multiple sequence alignment and phylogenetic analysis. Kindly suggest how to do it.
08 October 2017 1,814 4 View
I want to purify a 88 kDa bacterial secretory protein, which oligomerizes into different forms such as heptamers, octamers, dodecamers etc., for that I have buy Q Sepharose, Sepharose 6B phenyl...
07 October 2017 6,023 1 View
I'm analysis D and L glucose absorbance through UV spectroscopy and finding differences in absorbance for same concentration, what is the reason behind it.
26 September 2017 9,089 5 View
Nrf-2 is a modulators of different genes, so, why it is used as a marker for proteasome lysis analysis?
03 April 2017 9,263 1 View
We want to purify a 88 kDa bacterial protein from extracellular crude extract through gel permeation/size exclusion chromatography. I'm just beginner in this field and wonder about the basic...
27 December 2016 7,501 4 View
We want to purify a 88 kDa bacterial protein from extracellular extract through gel permeation chromatography. Kindly suggest suitable resin.
27 December 2016 5,299 6 View
I want to isolate secretory VacA protein from a broth culture. After salting out, I'm performing dialysis with 1X PBS (pH 7.5) and passing 1 ml of sample through self made 2 ml Sepharose 6B column...
14 December 2016 3,134 6 View
We are studying the effect of a toxin on cell membrane lipids which binds to receptors RPTP-alpha & beta but their localization in lipid rafts of cell membrane is not well characterized, so,...
07 September 2016 4,898 2 View
Suppose if mRNA is showing up regulation of a gene through qRT-PCR, then, is it necessary that the level of corresponding protein will also be high in western blot.
02 September 2016 1,472 4 View
Cryo-electron microscopy
13 April 2016 2,472 10 View
