Anti-Lamin A + C antibody [JoL3] - Nuclear Envelope Marker (ab4789)

We were wondering if you offer sample size aliquots of your antibodies, particularly anti Lamin A+C (cat # Ab4789)

Thank you,

ANSWER:

Thank you for contacting Abcam.

We do not supply samples sizes of our antibodies. Under our Abpromise we guarantee our products to work as stated on the datasheet and if for whatever reasons the antibody fails to work as stated then we will be happy to either to replace or refund the antibody.

If you wish to use an antibody in an untested application or species, then you can take advantage of our testing discount program, please see the link below for more information:

http://www.abcam.com/index.html?pageconfig=abpromise



If you would like to take advantage of the testing discount program or if there is anything else I can help you with, please let me know.

BATCH NUMBER 221038 ORDER NUMBER -- NOT SPECIFIED --

DESCRIPTION OF THE PROBLEM no signal

SAMPLE mouse brain progenitor cells and neurons; mouse embryonic and adult brain frozen sections

PRIMARY ANTIBODY mouse monoclonal to Lamin A+C 1:1, 1:10, 1:100, 1:500 were tried 4oC Overnight incubation or over weekend

DETECTION METHOD fluorescent microscope

POSITIVE AND NEGATIVE CONTROLS USED negative: without primary antibody Positive: no for primary. for secondary, we are able to detect b tubulin III (mouse IgG) and nestin (mouse IgG1).

ANTIBODY STORAGE CONDITIONS 4C, used immediately after receival.

FIXATION OF SAMPLE 4% paraformaldehyde

ANTIGEN RETRIEVAL no

PERMEABILIZATION STEP 1% or 3% triton in PBS for sections 1% triton or saponin for cells

BLOCKING CONDITIONS 10% NGS 30 min

SECONDARY ANTIBODY Alexa 488 goat anti mouse IgG or IgG1, 1:500 (Molecular Probe) RT 1 hr PBS 1%Triton wash

HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 3 HAVE YOU RUN A "NO PRIMARY" CONTROL? Yes DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes

WHAT STEPS HAVE YOU ALTERED? dilution, incubation time, permeablization

ANSWER:

Thank you for your enquiry. I am sorry to hear you are having a problem with ab4789.

I would like to suggest the following modifications to your protocol:

I was just looking at the data sheet, and unfortunately, this antibody has not been tested for tissue sections, it has only been tested for immunocytochemistry. We can not guarantee that this antibody would work in your mouse embryonic and adult brain frozen sections. As for the mouse progenitor cells and neurons, I would recommend changing your blocking step to 1% BSA. Have you tried to use the positive control listed on the data sheet, mouse myoblasts? Here is a link to our recommended ICC/IF protocol: http://www.abcam.com/assets/pdf/protocols/Immunocytochemistry%20(ICC)%20protocol.pdf.

If none of these tips help, I would be happy to offer a free of charge replacement or a refund if you have bought the antibody within the past 90 days.

Please let me know if this helps and do not hesitate to contact us for further advice.

BATCH NUMBER 176725 ORDER NUMBER SM635567

DESCRIPTION OF THE PROBLEM no band

SAMPLE Neuro-2A cells tried on (1) nuclear ectract (2)whole cell extract

PRIMARY ANTIBODY 1:50 diultions of ab in 5% milk PBS tween O/N incubate on rotator

DETECTION METHOD RCL Plus

POSITIVE AND NEGATIVE CONTROLS USED did siRNA of lamin A/C so shoud see a decreased if at all on the siRNA lamin A/C (Dharmacon) and use a negative control of siRNA (from Dharmacon)

ANTIBODY STORAGE CONDITIONS aliquot into 100ul each and store at -20 degrees Celsium

SAMPLE PREPARATION RIPA buffer for Whole cell extract

various salt buffers for nuclear extract-followed lab's protocol for it

AMOUNT OF PROTEIN LOADED 60ug for WCl extract 30ug for nuclear extract

ELECTROPHORESIS/GEL CONDITIONS reducing consition-used Invitrogen NUPAGE gel linear gradient 4-12% gel

TRANSFER AND BLOCKING CONDITIONS Block for 1 hour wiht 5% milk in PBS tween

SECONDARY ANTIBODY used Pierce's goat anti-mouse igG-HRP 1:2000 diultion in 5% milk PBS tween

HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 3 HAVE YOU RUN A "NO PRIMARY" CONTROL? Yes DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes

WHAT STEPS HAVE YOU ALTERED? add more anitbody to get 1;25 dilution and used 1:1000 dilution of secondary ab

ADDITIONAL NOTES film was clear -sis 1 hour exposure and still did not see the abnds for main A/C

ANSWER:

Thank you for the details that you have provided and I'm sorry to hear that you are experiencing difficulty with this antibody. Can you please tell me from which species your samples were obtained? Ab4789 reacts with Human, Mouse, Rat and Dinoflagellates and at a dilution of 1/50 should detect bands at approximately 70 kDa and 60 kDa. I just need to check a few things with you. Do you know that your samples transferred properly to the membrane? Is your secondary antibody working properly with other primary antibodies?

Thank you and I look forward to hearing from you.

Could you please tell me if the following antibodies with cross with porcine lamin: ab2559; ab4789; ab5090; ab8980. Thanks

ANSWER:

Thank you for your enquiry. All the information we have on species cross reactivity is specified on the datasheet, these are updated as soon as any new information is brought to our attention.

As far as we are aware, cross reactivity with pig has not yet been tested for use with ab2559, ab4789, ab5090 or ab8980. Should you decide to go ahead and purchase one of these products, please let us know how you get on by submitting an Abreview and in return we will award you 50 Abcam Points, which can be redeemed on a number of rewards (a further 100 points will be offered for an image).

Please let me know if you require any further assistance.

Hi,

Can I use this Ab (Lamin A+C) for detection of apoptosis because during apoptosis the 70kD lamin A is cleaved to a large and a small fragments. So I am wondering if this Ab can detect both full length and cleaved fragments (large and small)? Thank you.

ANSWER:

Thank you for your enquiry. It detects a C-terminal epitope so it will recognize the full length and I believe the smaller fragment. There is a paper in the European Journal of Cell Biology on this topic, please conduct a search for Broers J et al., 2002.