Sample preparation in PCR machine for SDS-PAGE?

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Query related to JC-1 mechanism

JC-1 dye is well known for using mitochondrial membrane potential disruption detection during apoptosis. JC-1 entry is from the cell membrane to the cytosol to the mitochondria and forms...

10 November 2012 5,049 15 View

Is it possible to use Alamar blue to test cell cytotoxicity at different time points with same sample ?

All these days we were using MTT for cell cytotoxicity. we purchased alamar blue from invitrogen newly, how can we use same 96 well plate reading after 1 day,2 day,3 days. Please share your...

08 September 2012 7,609 5 View

Is it possible to use Alamar blue to test cell cytotoxicity at different time points with the same sample ?

So far we've been using MTT for cell cytotoxicity. We purchased alamar blue from invitrogen newly: how can we use same 96 well plate reading after 1 day,2 day,3 days? As we know alamar blue is...

08 September 2012 9,113 3 View

Does an increase in caspase activity mean apoptosis should increase?

PI3k inhibitor(LY294002) and our test compound showed a 7 fold increase in caspase 3/7 activity . We further tested apoptosis by annexin V FITC+PI by flow cytometry. We found that in the PI3k...

07 August 2012 4,198 14 View

My HepG2 cell line is growing very slowly, why?

I am trying to grow hepatic cell lines like HepG2, HUH-7 and ,Hep-3b for anticancer work. I am growing all in RPMI-1640 + 10%FBS + 1X Ab/AM. HUH-7, and hep-3b are growing very nicely but HepG2...

03 April 2012 4,414 10 View

how to calculate IC50 using sigma plot software.

I do MTT assay to see the cell viability upon drug treatments.we have sigma plot.10 software programme.could anyone tell how to calculate IC-50 using that.

01 February 2012 9,410 5 View

Solvent for partially purified product?????????

We have some plant product partially purified powdered form,i need to check anticancer activitiy of that.iam dissolving product in Methanol and further dissolving in growth medium for MTT...

11 December 2011 6,573 3 View

Cleaved caspases detection

I would like to detect cleaved caspases (17,19 kd) so Is 15% SDS PAGE ok for this? running SDS PAGE 25mA for 1hr and western transfer conditions i use 100V for 1hr. and any suggestions during...

11 December 2011 3,747 1 View

Does anyone know what might be causing the smeared bands on my western blot?

I got these smeared bands quite often lately. We typically run the gel at 140V with a 10-12% gel and do a wet transfer at 220 mA for 1.5 hr in cold room. We also noticed some dirty spots/dots (see...

10 August 2024 7,480 3 View

What will be the best way to test NFkb activation via western blot?

Hi, I have lysate from human macrophages, and I wonder what will be the best way to check the NFKB activation in those cells. I was thinking p-p65 or p-ikkb./ Is that make sense?

18 July 2024 5,214 4 View

How to detect phosphorylation on Western Blots?

Having trouble detecting phosphorylated HSL. Any tips for how to prevent de-phosphorylation events or detect phosphorylation?

15 July 2024 2,055 3 View

Trouble finding an anti-Digoxigenin antibody to perform Southern Blot technique?

I'm currently using Roche anti-DIG-AKP Fab fragments to perform Southern Blot assays, but this reagent is delayed since november last year. It's anybody facing the same problem? If so, how are you...

15 July 2024 6,668 0 View

Is there anyway to distinguish two proteins of same molecular weight?

I know mass spectrometry will work. But, is there any simple and straightforward method to distinguish between the two proteins of the same molecular weight? They are cryptochromes and...

14 July 2024 9,324 2 View

What would you do next?

Studying a specific monogenetic disease based on mutations in different individual genes. I try to recreate the disease phenotypes of donors in vitro. I knocked out a gene x with above 90%...

12 July 2024 6,313 0 View

Loading control for western blot to detect Histone H1.2?

Hello, I isolated total histones successfully from the skin primary cells by acid extraction. I wanted to detect histone H1.2 expression in different conditions by western blot. Can some one...

10 July 2024 9,165 1 View

Personal recommendations for GAPDH antibody?

Hello! I am planning a western blot experiment, and I am looking for GAPDH antibodies to order. In the past, I have experienced a lot of hit-or-miss when ordering antibodies based on just the...

07 July 2024 1,685 0 View

How can I solve this problem with my western blot?

I have this problem with my western blot for the ABCG2 antibody on nitrocellulose membrane. Help please

02 July 2024 8,715 2 View

How to troubleshoot error E1 on Biorad Powerpac Basic despite using sufficient and fresh buffer?

I have used this BIorad Powerpac Basic for 4 years now, before which it was used for another 5-6 years. Generally we addressed the E1 error by compensating with fresh buffer and/or filling the...

01 July 2024 7,104 1 View