How much Fe-ions are in cancer cells?

Hi, I have a question for someone with experience in cancer cell metabolism. I wonder how much iron ions are in cultured cells [free or other wise available for haeme synethesis] using 10% FBS. I...

22 May 2018 4,842 4 View

Why does my standard curve not generate expected concentrations in the back calculation?

I am setting up a direct ELISA and have a standard curve which I fit to a 5-PL curve. When I try to calculate the concentrations of the points in the standard curve using the obtained 5-PL curve I...

12 March 2018 1,410 2 View

Does anyone have Hudson and Levasseur( 2002): Supporting foster parents: Caring voices.

10 July 2017 9,532 6 View

What is a good software for curve fitting?

I am looking for a good software for fitting curves and would like to hear what software other people recommend. I want to be able to fit 4- and 5- parameter logistic curves and linear regression....

09 April 2014 2,980 49 View

Optimal cell (HEK-293) density for transient transfection and IgG expression

I am wondering what your opinion is on what cell density is optimal for expression of antibodies by transient transfection of suspension adapted HEK-293 cells in batch culture using PEI as...

25 February 2013 7,774 2 View

Protein concentration using Amicon Ultra spin column, adsorption on membrane?

I am trying to concentrate a protein using Amicon Ultra centrifugal device. I have done this several times before without any problems. This time I had a larger volume of my sample so I had to...

15 October 2012 8,778 34 View

Protocol for internalization of antibody?

I want to set up an assay to detect internalization of an antibody in HEK cells. I want the assay to be ELISA based since I don't have access to a FACS machine or a confocal microscope. Does...

07 October 2012 741 20 View

Imidazol concentration for His-Trap purification

I am purifying a protein from a cell supernatant with a His-trap column from GE. I get quite low yield and it seems as if the protein doesn't bind very well since I can obtain protein when...

05 September 2012 4,507 43 View

IgY purification with low g centrifugation?

I want to purify IgY from eggs using a PEG based protocol since this is cheap and fairly simple, compared to amoniumsulphate methods. I have only found protocols where one has to centrifuge at...

31 July 2012 1,194 7 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

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

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

Are exogenous molecules transferred between human cells during doubling ?

Hello RG community, I wonderer whether exogenous molecules (e.g. reporter plasmids, miRNAs or other artificial molecules) are transferred between cultured human cells during their doubling ? So,...

21 February 2021 6,633 3 View

Which transfection reagent is the best for plasmid transfection of MDA-MB-231 cell line, Lipofectamine based or other reagent suggestion?

I try to transfect MDA MB 231 cell line with a plasmid by PEI, unfortunately, the rate of transfection is low, and now I wanna find and order the best transfection reagent for this cell line and...

19 February 2021 828 6 View