Any advice on checking for green fluorescent protein in one of the cancer lines in FACS canto?

More Komal k kumar j's questions See All

I am confusing with antibody dilutions can any one help me ?

I have purified an antibody the final concentration is 0.42mg/ml. i want to titrate antibody, so that i can pick up 0.5 microgram/ml to 5microgram/ml concentration in 100 microL staining medium. i...

31 December 2014 2,652 2 View

Can anyone tell me how to save Human Leukemic cells after treating with drugs before preparing peptides for Mass spectra?

Can anyone tell me how to save the cells (Human Leukemic cell lines) after treating with drugs before preparing proteins for Mass spectra? i have other things to do so i have planned to save the...

11 December 2014 6,353 2 View

Can anyone tell me the suitable stimulation conditions to stimulate pStat1, pStat3, pStat4, p38 and pAkt?

Hi guys. Can anyone tell me the suitable stimulation conditions to stimulate pStat1, pStat3, pStat4, p38 and pAkt. I am working with mouse spleen, bone marrow and murine cell line BaF3. I am...

10 November 2014 8,197 2 View

Can anybody help with multicolour flow cytometry analysis?

I performed multicolour analysis in flow cytometry. I stained for FITC, PercpCy5, PercpCy7, PE and PB on cancer cells. I compensated using Beads and used FMos for gating population, but when I run...

03 April 2014 4,150 14 View

Phosphoflow: I am working on cancer cell lines looking for phosphoproteins by flow cytometry. Any advice?

In flow cytometry single histogram of a single phosphoprotein splitted into two peaks. Even that is a stable cancer cell line. Why is this?

11 December 2013 647 0 View

Why does a single histogram sometimes split into two peaks for a single phosphoprotein in flow cytometry?

I am working on phosphoflow for specific proteins. In the histogram sometimes a single histogram split into two peaks for single phosphoprotein, but sometime for the same protein that peak...

11 December 2013 5,684 5 View

Green fluorescent proteins peak is too high at FITC channel for all the samples - any suggestions?

I added PI to the cell line that I am interested to exclude dead cells. Then run the experiment in FACS canto. But GFP is sowing the peak at 10 to the power of 5 in FITC channel for all the...

11 December 2013 5,469 5 View

What should nonphosphorylated and phosphorylated proteins looks like in in the histogram?

I am working on phosphoflow in FACS and have stained the cells with surface markers and intercellular markers. I got the FSC and SCC results and histogram but I am confused about how to interpret...

10 November 2013 10,032 4 View

Why does my protein refolded to beta sheet during thermal denaturation analysis?

Hi! So i attempted to understand a novel protein behavior towards heat application by analyzing its secondary structure change. I subjected the protein to a thermal denaturation analysis using...

06 August 2024 1,989 3 View

How to determine positive-stained cells in FACS? Use isotype or unstained control?

To compare positive and negative cell populations in flow cytometry, should I compare unstained cells with antibody stained cells? Or with the isotype control? Most papers show comparison with...

06 August 2024 6,728 6 View

Why do we equate male and female arousal?

Women, on the other hand, can become physically aroused (increased blood flow in the reproductive organs) without becoming psychologically aroused even in the slightest. (Robert Weiss)

05 August 2024 9,537 2 View

Could dyes amplify the spectrum of light to a specific wavelength?

I am interested to know the behavior of dyes toward light. Specifically, Blue dyes re-emit the spectrum, especially from the green zone (known as principal in LED lamps, and blue dyes are known...

05 August 2024 3,290 1 View

What is the problem with these tissue culture plants?

All plants are green but some of these plants becomes yellow. I did not found any reason. Please help me to find out the real problem.

01 August 2024 589 4 View

How to make a Lateral flow control line more uniform?

We have a lateral flow test on Sartorius 140 NC. The conjugate is gold-monoclonal antibody. Using different control lines (GAM, protein A, protein G) we get a very strong leading edge with reduced...

31 July 2024 4,510 3 View

Transfection in HEK293T cells?

Dear All, I am trying to transfect a pCDNA3.1 vector containing my gene of interest. The purpose is to figure out the localization of the protein of interest. I have fused the protein with GFP on...

31 July 2024 9,892 4 View

How to retain the a GFP tagged gene expression in stable cell line?

Hello , I established a stable cell line expressing GFP tagged to a centrosomal gene having G418 drug selection marker. I validated the stable line by IFA and Western blotting, results are fine....

29 July 2024 5,007 0 View

Suggestions for the Annexin V Positive Control ?

Hello everyone, I am currently using washed human platelets to stain Annexin V as a procoagulant marker. Additionally, I am staining with PerCP-CD61 to identify platelet cells. So far, I have...

29 July 2024 8,624 1 View

What is the relationship between protein structure and N or C terminal tagging choosing?

I want to do 2,3-butanediol dehydrogenase(BDH) enzyme purification to confirm its activity for 2,3-butanediol. Before that, I need to confirm which N or C terminal tagging is better for enzyme...

28 July 2024 366 3 View

Peter D Pioli

Hi Komal,

Use the FITC channel.

Pete

Dmitry B. Kazansky

Peter is right. FITC (FL1) will be good.

Komal k kumar j

Thanku peter and dmitry

Susan Christo

Depending on how the configuration was set, the Canto should have GFP from its dropdown menu? Actually, maybe that's the Canto II. Anyway, I echo Peter and Dmitry - FITC/FL1!

Ok thank u susan christo

Farid Ahmed

Use FL-1, or FITC channel (530/30 filter) on Canto

Thank u farid

Rahul Sharma

unanimous suggestion to use ....FITC/FL1 channel... :) :)

Yes