DCF_DA fluorescense -ROS detection in attached cells

Hello everyone Recently I have been working on ROS production studies using the dye DCF-DA fluorochrome. I came across idea that trypsinization also increases the ROS production in cells,...

03 April 2011 2,536 22 View

Western Blot-Bcl-2 Total Santa Cruz polyclonal antibody- ghost bands numerous

Hello everyone, I have started doing western blot for some months but then with the new antibody from Santacruz for Bcl-2 I got lots of ghost bands.... I have tried reducing d concentration of...

02 March 2011 6,311 6 View

Phosphorylation studies-how effective is using animal serum for blocking? TBST & PBST any significant difference?

I was doing a phosphorylation study of Tau using tau-1 antibody from Millipore, I am having trouble getting bands... on contacting the company they asked me to change the blocking solution to the...

02 March 2011 6,930 6 View

DNA ladder apoptosis-no ladder formation

Hello Everyone, I once tried performing the DNA ladder test as a measure for apoptosis. I isolated the DNA using a kit and also quantified it, it was ok back then even purity seemed ok. However...

01 February 2011 5,043 19 View

Needed some help with flow cytometry

Hello everyone, I just started with the flow cytometry and since I was doing it for the first time, I planned to move on with single PI staining for detection of dead cells-either late apoptosis...

10 November 2010 2,861 18 View

Markers of Oxidative damage-PCC..Protein turnover

Protein carbonyl content is the mostly employed marker for oxidative damage. Protein turnover seems to be a old parameter used for the similar pupose. Morphological damage is a good indicator but...

09 October 2010 2,845 3 View

N Acetyl cysteine and MTT--incompatible or am I only having problems

I recently tried using NAC on some cell line and wanted to check the viability of cells using MTT. I found that as soon as i add MTT to wells containing NAC it turns bluish black......some kin dof...

09 October 2010 3,936 1 View

Nuclear Staining methods--DAPI and Hoechst- Apoptosis....TUNEL......

Nuclear staining methods DAPI and Hoechst are both minor groove binders intercalating the AT regions. In case of apoptosis the cells may show ssDNA strand breaks as well as dsDNA strand breaks-...

09 October 2010 8,770 5 View

N Acetyl cysteine and MTT--incompatible or am I only having problems

I recently tried using NAC on some cell line and wanted to check the viability of cells using MTT. I found that as soon as i add MTT to wells containing NAC it turns bluish black instantly...

09 October 2010 2,174 4 View

Wanted to ask my senior researchers about quality of antibodies from Anaspec

Hello I am thinking about buying my antibodies from Anaspec, california USA, have any of you used their antibodies, just wanted to be sure of the quality of the product that i shall be purchasing...

01 February 2010 4,823 0 View

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02 March 2021 3,060 3 View

Issues with Permeabilization during Flow Cytometry?

Hello Everyone. Currently I am working to characterize macrophages in the myocardium after ischemia-reperfusion injury in rats. Due to the low total cell number isolated from rat hearts I can...

01 March 2021 3,867 3 View

Which cell line should we use to perform biological dosimetry experiments?

We are preparing some experiments based on irradiating cells under different conditions in order to evaluate the effects in terms of DNA damage, genetic expression, etc. As our project is...

01 March 2021 3,355 3 View

Is it possible for two human neuronal cell lines to have different gene sequence for the same protein?

I am looking at the ATP1A2 (Sodium/Potassium ATPase alpha subunit 2) in two human neuronal cell lines. Expression levels of this protein seems to be almost equal when detected by one antibody....

01 March 2021 3,607 3 View

Is CD44 degraded in lysosome together with hyaluronan after CD44 mediated endocytosis of hyaluronic acid?

Also when RHAMM binds hyaluronic acid, they get internalized, will RHAMM also be degraded? Or both CD44 and RHAMM will be transferred back to the cell membrane? Asking for breast cancer cell line...

01 March 2021 8,169 2 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

What could be some reasons why unstimulated monocytes are getting activated?

Im doing PBMC isolation -> CD14+ enrichment using magnetic beads -> stimulation setup. My negative control is just cells in cRPMI but they seem to get activated over and over again.

28 February 2021 7,883 3 View

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28 February 2021 9,936 2 View

When is it unnecessary to change organisational culture?

As a former management practitioner, I recall a generally accepted view that changing organisational culture is difficult, so you should only attempt it if the organisational change that you are...

27 February 2021 2,595 8 View

Does RNAiMAX transfection reagent work in serum free medium?

I diluted siRNA and RNAiMAX in opti-MEM and added to the cells which they were in the growth medium. Is it a right way? or should I culture cells in the opti-MEM medium for a while and not in...

26 February 2021 10,041 3 View