Does anyone know properly analyze CLS stuctures? What features should I look for (eg. small adipocytes surrounded by cells)?

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Multiple sgRNA cloning into pX333 plasmid using BsaI and BbsI restriction enzyme site?

Currently, I am using a plasmid containing two gRNA cloning site. The restriction enzymes used for cloning gRNAs into this plasmid, pX333, are BsaI and BbsI. I digested the pX333 plasmid using...

09 October 2019 2,895 6 View

How do I design primers specific to transfected plasmid and not genomic DNA?

I recently introduced a Lenti-Puro Guide vector containing a specific sgRNA sequence into a cell line. I would like to sequence the ddCas9-sgRNA cell lines to confirm that each sgRNA is accurate.

01 February 2019 5,439 3 View

Has anyone uses a 4in1 cholesterol tester on research mice?

Hi all, Has anyone used a 4in1 cholesterol tester on research mice? We have used the One Touch handheld glucose monitoring system to measure glucose on high fed diet fed mice. However, has anyone...

09 October 2016 3,948 2 View

From a budding scientific illustrator and PhD student - Are you in need of a scientific illustrator?

Are you in need of a scientific illustrator for creating illustrations (cellular and molecular mechanisms, structures, etc) to compliment a primary paper or review article? If so, please let me...

07 August 2015 5,479 0 View

Should I serum starve primary hepatocytes if I wish to probe for pAMPK using westen blotting?

I am using stimulated primary hepatocytes to detect alterations in AMPK activity. However, a paper published that serum starvation can increase AMPK activity in different cell types. Although,...

07 August 2015 8,015 3 View

Does anyone know of easily transfected mouse cell lines?

I would like to assess the editing efficiency of designed guide RNAs by transfection prior to establishing a stable KO cell line (via lentiviral infection). HEK293T cells are very efficient when...

01 January 1970 6,500 2 View

Do I need specific antibodies to stain for a fusion protein?

I am staining some brain sections stored in cryoprotectant that express a Histone H2B- GFP fusion protein that were generated ~10 years ago. I know I need to enhance signal with an anti-GFP...

07 August 2024 5,338 2 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

Can I multiplex a mouse monoclonal and rat primary for IHC (mouse brain tissue)?

Hi all, I was just wondering if anyone has experience with multiplexing a mouse monoclonal primary and a rat primary. I'm trying to multiplex by incubating them in the same well but was told by a...

06 August 2024 9,710 1 View

Senescence-associated beta galactosidase staining is False Positive in control group?

Hi guys If anyone is currently working on aging cells, you guys would like to give me some advice. I'm testing against biomarker (SA-beta-Gal), I encountered a false positive in the control group...

02 August 2024 6,735 1 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

How can I get clearer background for my western blot?

I am running a western on HIV envelope protein. The size is around 50Kda and I am using rat anti-HA for the first stain and a second stain of mouse anti-CA. Primary antibody dilution is 1: 1000...

29 July 2024 6,484 5 View

How many events should be counted for the analysis of apoptosis with nuclear staining with Hoechst 33342 whit microscopy ?

Greeting How many events should be counted for the analysis of apoptosis with nuclear staining with Hoechst 33342 whit microscopy ? Are 100 events count sufficient and three repetitions for...

25 July 2024 7,520 0 View

Ideal tissue thickness for multiple immunolabelling and confocal microscopy?

Hi there! I'm hoping to to look at co-localisation of cytokines and markers of different brain cell types (e.g., neurons, astrocytes, microglia) in juvenile rat brains (postnatal day 28) that have...

23 July 2024 2,844 1 View

What are those black/hollow structures?

Hello, I recently stained my cells with an Arf6 antibody. The healthy control looks pretty normal, while the disease group cells have abnormally large black structures in the cell body. Has anyone...

21 July 2024 1,044 1 View

Fixation Problem with cancer cells?

Hello everyone, I work with polyacrylamide hydrogels, which I fictionalise with sulfo-sanpa and collagen I to then plate lung cancer cells. The problem is that I lose a significant amount of...

19 July 2024 3,906 0 View