Can I use blocking peptide as positive control in Western blot?

I'm just curious if blocking peptides can be used in this fashion? Does the conjugation of the blocking peptide with the primary antibody stop the immunoreactivity with say a flouro-tagged...

31 December 2015 5,556 5 View

MDA-MB-231 advice?

I'm looking to do a time course realtime-glo vitality assay on MDA-MB-231 in a 96-well plate. I plan on doing 4 different seeding concentrations to assess linearity of signal, however there is...

05 June 2015 1,797 4 View

How can I perform SiRNA concentration scaling? by nM or pmol per num of cell / surface area?

31 December 2014 4,760 6 View

How can I explain great siRNA transfection efficiency but no knockdown with validated positive controls?

I'm performing reverse transfection on SH-SY5Y using Viromer Green, and I get wonderful transfection efficiency (90+%). See picture (RFP labeled siRNA fluorescence overlapped on phase contrast...

10 November 2014 7,460 7 View

Can anybody recommend how much Blasticidin should be used for selection of SH-SY5Y?

We are electroporating a pcDNA6.2 plasmid contain miR RNAi constructs into SH-SY5Y. I plan on selecting these cells with blasticidin, and the literature suggests closer to the 10ug/ml mark for...

09 October 2014 6,422 3 View

Is anyone familiar with electroporation?

I thoroughly pipette my cells prior to electroporating so they should form a monolayer when plated. Though every time I electroporate, they clump. I'm not entirely sure why. When I passage, they...

08 September 2014 6,324 3 View

Shsy5y immunostaine troubleshooting

cells plated

07 August 2014 2,161 0 View

What is a good number of SH-SY5Y to freeze down for long term storage between 1-12mo?

What is a good number of SH-SY5Y to freeze down for long term storage between 1-12mo? Ideally, I'd like to initially plate in something larger like a t75 or t150, though I'm not sure if it is...

06 July 2014 4,719 2 View

What is the best method to carry out SH-SY5Y differentiation and transfection?

How do people typically go about electroporating differentiated SH-SY5Y? I was thinking that you would have to electroporate them first then recover in media supplemented with retinoic acid. Would...

06 July 2014 1,714 1 View

Can anyone share their knowledge on cell penetrating proteins?

06 July 2014 1,383 2 View

In PRIMER, is it possible to downweight certain taxa in multivariate community datasets to reflect uncertainty around identification??

I have a dataset with about 80 different species. As usual, some species are very easy to identify with certainty whereas others are more difficult, which means that I am less certain of my...

03 March 2021 8,066 4 View

Is There Any Feasible Method To Test Fluorescent Compounds Other Than Fluorescence Spectrometers ??

Is There Any Feasible Method To Test The Efficiency Of Fluorescent Compounds Other Than UV Spectrometers ? Suggestions Would Be Appreciated !

02 March 2021 5,785 3 View

PACYC PCR not working after several attempts. Would anyone have any suggestions?

So, I have been trying to run a pACYC PCR which will be used later on for a Gibson Assembly. However the PCR is not working. I have already tried gradient PCR and changing extension time; however...

02 March 2021 1,146 2 View

How to calculate the annealing temperature of a primer pair?

I have to amplify a gene and my primers just reached. The Tm for Forward primer is 64.2, and that of reverse primer is 65.5. Can some one suggest how to get the best annealing temperature? Thanks...

01 March 2021 360 7 View

No title

Does anyone have the experience of using Taq Man probes in the QIAGEN Rotar- Gene qPCR machine?

01 March 2021 5,311 1 View

Any research papers for extraction of nutrient deprivation on macroalgae? Analysed by HPLC-UV and GC/MS. Possibly from macroalgae genus Rhizoclonium?

I am doing my thesis on the nutrient deprivation on the macroalgae Rhizoclonium sp. I will be analysing the levels over a period of roughly 5 days by HPLC-UV and GC/MS. I will be using seawater as...

28 February 2021 6,477 1 View

Anybody with an idea on how do design specific primers for detecting tomato resistance genes tm-1, TM-2 and Tm2^2?

I am trying to identify these 3 genes among some tomato cultivar collections and after aligning some sequences from NCBI, I couldn't find unique sequences to target for specific primers. There...

28 February 2021 606 3 View

What is the ideal efficiency for a qPCR?

hello everyone, I need to do standard curves for my qPCR, what is the ideal efficiency range? I tried a primer (Mglu2 receptor) that gave an efficiency of 90.2%. Is it accepted?

28 February 2021 1,254 3 View

MiRNA qpcr problem?

Dear All, mirna primer showing some problem in the melting curve? any idea why? As attached is the melting curve. The forward sequence is obtained from miRBase and reverse primer is universal.

28 February 2021 5,008 4 View

Calculating volume of DNA required from DNA concentration?

Hi I am a bit confused. They are asking me to find out the volume of DNA required in ul (a total of 30-100 ng for genomic DNA) from the DNA concentration in the nanodrop reading which was 404.8...

26 February 2021 5,029 2 View