Why stop the reaction in ice for the fluorescent probe FDA (fluorescein diacetate)?

Hi, In literature I noticed that who stains haemocytes with FDA for 30 min then stop the reaction in ice while this does not apply for 15 min incubation time (or maybe authors did not write this...

08 February 2021 3,366 3 View

Which is the importance of fixing (e.g. bivalvia) haemocytes and use anti-clotting solutions in flow cytometry?

I am trying to optimize a flow cytometry method to characterize mytilus spp. haemocytes. In literature I found that someone, to avoid clotting, use anti-clotting solutions or dilute and fix the...

05 November 2020 2,522 4 View

Has any of you encountered issues with protein production in a E.coli cell-free protein expression system?

I'm having trouble synthesizing a mollusk 173 aminoacids protein of unknown function in a E.coli cell-free protein expression system. The positive control works well and the plasmid has been...

01 October 2019 8,829 7 View

How I could set up a lipoplexe-based transfection protocol and compare data with commercial Viromer Red transfection reagent?

I am new in the field of transfection and I'm having a lot of problems in setting up the protocol with LIPOPLEXES in order to compare data previously obtained using Viromer Red protocol on THP-1...

13 November 2018 3,545 1 View

Does anybody know a reasonable (in prize) company where to buy an environmental chamber to adapt a confocal microscope for live cell imaging?

We are interested in adapting a Confocal microscope for long-term live imaging, and for this reason we are trying to get a small environmental chamber at a reasonable prize. It could be enough...

16 May 2017 5,437 2 View

How to store a purified protein before a kinase assay?

23 September 2013 8,697 1 View

Is it possible to position the HA taq not at the extreme end of a protein and visualize the taq-protein by Immunocytochemistry?

06 June 2013 5,942 9 View

What are the best antibody markers to detect and quantify the circulating endothelial cells and progenitors by flow cytometry?

CD45/CD31/CD34/KDR or CD45/CD31/CD146/CD133? Or both or neither?

18 February 2013 1,958 2 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

Can someone suggest a way to prepare slides to observe under confocal microscope for observing fluorescent tagged proteins??

I am growing the cells on coverslips and transfecting the fluorescent tagged protein directly on them. Then i want to observe these cells under confocal microscope. Currently i am just using...

01 March 2021 6,142 3 View

Problem with shRNA transfection. Why are only two plates getting contaminated instead of all?

I transfected my LNCaP-WT cells with 3 shRNA plus their NTC two weeks ago and split two puromycin selected cell plates on Friday last week(Feb 26). I checked for GFP in the cells, and they all...

28 February 2021 4,949 3 View

Is there anyone who experienced with cytotoxic effect of Hoescht 33342 dye?

Is Hoechst toxic to cells? In general, it is known as non-toxic. I am working with dsRED MDA-MB-231 cell line. I applied it for my PDMS-based microfluidic platform, for this reason, I incubated...

26 February 2021 9,119 3 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

How do I transfect a large plasmid into PC12 cells with nerve growth factor (NGF) receptor?

I have a large 16 kb plasmid, which I need to transfect into PC12 cells. Lipofectamin 2000 didn't work and with GFP alone the transfection rate is very low. I also tried the neon invitrogen...

25 February 2021 6,635 5 View

What is a good reagent for lentiviral plasmid transfection in HeLa cells?

I have been using Lipofectamine 2000 but was hoping there may be a better protocol or one for something like Lipofectamine PLUS reagent.

24 February 2021 1,633 4 View

Why the cells die after stable transfection?

I tried stable transfection in the U87 cell line with the sgRNA-Cas9 All-in-One lentivector. Cells were cultured in a 6-well plate, 4ug plasmid was transfected after 24hs when the cell confluence...

22 February 2021 2,864 4 View

Does anyone have any ideas why drug-resistant malaria is increasing in Southeast Asia even when those drugs are no longer used?

When dealing with bacterial toxins, there are three varieties of microbes that develop in your petri dish: toxic, resistant, and susceptible. Initially, there are only toxic and vulnerable...

21 February 2021 6,084 5 View

Is there a method for evaluating nanoparticle adhesion to epithelial tissue in vitro?

I want to measure how well nanoparticles can transfect cells based on NP adhesion properties. In the literature I find people do it in animal studies, and epithelial adhesion helps in transfection...

21 February 2021 7,791 3 View