Anti-MMP17 antibody [EP1270Y] (ab51075)

Please see customers response below:

With regards to the antigen retrieval issue, we have in fact already tried several different durations and even at times not using antigen retrieval at all, triton-X was only used in some runs. All of our other antibodies are ok when used in conjunction with our IHC kit.

At this point, i think we would prefer a credit note and use it to buy another MMP17 (ab39028) from Abcam. Just wondering what the process is for that? can we exchange for a different product without being issued a credit note?

Looking forward for your response.

Kind regards,

ANSWER:

Thank you for your message and for providing this further information.

I am sorry there are no further suggestions to help improve the results on this occasion. I appreciate the time you have spent on these experiments and would be pleased to arrange a free of charge replacement or credit note in compensation. Alternatively, I am happy to provide a free of charge vial of a different antibody. Please let me know which one you would like to receive.

I look forward to hearing from you with details of how you would like to proceed.

Product code: 51075
Lot number: GR19885-6
Inquiry: Please see technical enquiry form attached.

Immunocytochemistry/Immunohistochemistry/ Immunofluorescence
ANTIBODY DETAILS
Antibody Code: EP1270Y (ab51075)
Batch Number: GR19885-6
GENERAL INFORMATION
Antibody Storage Conditions (temperature/reconstitution etc)

Arrived in liquid form from manufacturer

Description of the Problem (high background, low signal, non-specific staining etc)

No staining

Sample (Species/Tissue/Cell Type/Cell Line etc)

Human placental tissue

Fixation of Sample (Ethanol/Methanol/Acetone/Paraformaldehyde/Other/Duration)

We have attempted on both formaldehyde and paraformaldehyde fixed tissue (duration of fixation is usually overnight at 4 degrees

Antigen Retrieval (Enzymatic method, Heat mediated technique etc)

We have attempted enzymatic (Trypsin, using Invitrogen’s Digest All Kit), and heating sections in citrate buffer as well as EDTA antigen retrieval buffer for 15-20minutes at high & low power.

Permeabilization step

We have tried Triton-X as well as

Blocking Conditions (Buffer/time period, Blocking agent)

We use Protein Block (DAKO) for 10minutes

Primary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, wash step)

Abcam Antibody/Rabbit/1%BSA,PBS/1:10, 1:25, 1:50/1-2hr at RT or 37 degrees, overnight at 4 degrees attempted as well/washed with either PBS or PBST after primary antibody
Secondary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, wash step)

Invitrogen: Superpicture 3rd gen IHC kit/Universal including rabbit, mouse, rat/ready to use/10-30 minutes/wash with PBS

Detection Method

DAB

Positive and negative controls used (please specify)

Placental tissue is a positive control for MMP17. Placental tissue has been shown to express MMP17 by both Western Blot and Real-Time PCR methods.

OPTIMIZATION ATTEMPTS (PROBLEM SOLVING)
How many times have you tried the IHC?

30+ sections have been used to optimize the antibody, we have tried everything
Have you run a “No Primary” control? Yes (Delete one)
Do you obtain the same results every time? Yes (Delete one)
What steps have you altered?

Tried altering incubation time, antibody concentration, antigen retrieval methods (duration and solution)

Additional Notes

The antibody does not work at the highest concentration recommended by Abcam (which was 1:50). We have tried even higher concentration without success.

ANSWER:

Thank you for taking the time to complete our questionnaire and contact us. I am sorry to hear you have had difficulty obtaining satisfactory results from this antibody.

The details you have kindly provided will provide us with vital information for our monitoring of product quality. I would like to reassure you that this antibody is tested and covered by our guarantee for IHC-P and human samples.

I appreciate the time you have spent in the laboratory and understand your concerns. It is regrettable the results have not been successful. Reviewing the details, I am sorry there not many tips to provide on this occasion to help improve the results. However, I can recommend you may like to consider trying the following options and considerations:

1. I can suggest to try less time for heat mediated antigen retrieval? This can sometimes require some optimization, and less time is sometimes better than more time. For example, try 5 minutes.

2. As MMP17 is a cell membrane protein, we would not recommend to permeabilize in Triton. Triton is a strong detergent which can disrupt membrane proteins. It can sometimes significantly affect the staining of membrane targets.

3. Is the IHC kit working well with other primary antibodies?

Alternatively, or if these tips do not work, I am pleased to offer you a free of charge replacement or credit note in compensation if the item has been purchased in the 6 month guarantee period. In this case I would appreciate if you are able to confirm the order number.

Thank you for your cooperation. I look forward to hearing from you with the results of the suggestions details of how you would like to proceed.

Dear technical support



Would you be able to advise me on the concentration as per ab51075 (anti-MMP17). The datasheet sheet does not specify the concentration.



Looking forward to your response.



In the meantime, have a nice weekend J



Kind regards,

ANSWER:

Thank you for your inquiry.

The concentration of ab51075 is lot dependent. The lot that we have in stock at the moment and of the last vial you receivedhas a concentration of 0.224 mg/mL.

I also would like to comment on the seemingly low concentration:

The IgG produced by rabbits have greater affinity and greater sensitivity than mouse IgG (about 100-1000 fold) and at the same time the total amount of IgG produced by the rabbit hybridomas is generally lower than in mouse hybridomas.

Many 'veteran' antibody users are generally familiar with the IgG amount provided from mouse antibodies and can judge how many reactions they may get or need to use based on concentration. Unfortunately, this experience is not transferable to rabbit monoclonal antibodies because the concentration scales are generally very different.

Since the specificity and sensitivity is much higher with rabbit monoclonal antibodies, less IgG is required per reaction.

I would like to reassure you that while the amount of IgG may not be comparable to mouse monoclonal antibodies, customers are receiving a comparable number of reactions/tests/blots/stainings per tube.

I hope this information is helpful. Please do not hesitate to contact me with any further questions in this matter and have a good weekend too.

Immunogen von ab51075?

ANSWER:

Vielen Dank für Ihren Anruf gestern.

Ich habe jetzt genauere Information zum Immunogen erhalten:

Es liegt zwischen den Aminosäuren AA190 und 230 (SwissProt ID Q9ULZ9).

Ich hoffe, diese Information hilft Ihnen herauszufinden ob die zusätzliche Bande auf dem WB dem "ge-sheddeten" Teilstück des Proteins entspricht.

 Hi,   I’m wondering how long you suggest an antibody lasts for – I purchased several antibodies about eight months ago and none of them came with an expiry date, the info sheets just say “store at -20 long term”.  These antibodies are:   AB38971 Rabbit Polyclonal to MMP14 - Hinge region Lot # GR12166 AB56308 Mouse Monoclonal [MM0028-5D11] to MMP15 Lot #GR40525 AB73877 Rabbit Polyclonal to MMP16 Lot # GR30734 AB51075 Rabbit Monoclonal [EP1270Y] to MMP17 Lot # GR19885 AB39031 Rabbit Polyclonal to MMP25 - Hinge region Lot # 275516   Thank you,      

ANSWER:

Thank you for your enquiry.

We do not have set expiration dates for our products. Most antibodies are stable and can last for anywhere from a few months to several years if stored properly, so we strongly recommend that you follow the storage instructions on the datasheet for the antibody you purchased. These conditions will vary among our antibodies, therefore, it is important to verify the storage conditions for each of our products when you receive them. We guarantee all of our products to work for at least six month from the date of purchase when stored correctly.

For more information on antibody storage and stability, please visit our Antibody Storage Guide on our Protocols and Troubleshooting Tips page (http://www.abcam.com/index.html?pageconfig=popular_protocols).

I hope this information helps. Please do not hesitate to contact us if you need anything further.