Anti-D Serine antibody (ab6472)

I saw the following question on your website. Can your D-serine antibody specifically recognize D-serine. How would you set up the ELISA assay ? Do you have more detailed information on this

Question 1 Wednesday 02-June-2004 Beginning of February 2004 I received D-serine Antibodies ab6472 which I would like to use to establish an D-serine ELISA for following D-serine blood level in patients with liver transplantation. System: Kompetitive ELISA Plate: Exiqon Coating: BSA-G-D-serine, BSA-G-L-Serine, OVA-G-D-serine, OVA-G-L-serine, over night, extensive washing with phoshate puffer Incubation of anti -D-serine antibodies (ab6472) for 3 hours, extensive washing with phoshate puffer, Tested dilution 1:2500, 1:5000, 1:10000 Incubation of anti-Rabbit-Antibodies conjugateted to HRP (used as Standard for other applications), 1 hour, extensive washing with phoshate puffer Addition of TMB (15 min.) and stopp-solution. Signals (Extinction)): For all haptens: BSA-G-D-serine, BSA-G-L-Serine, OVA-G-D-serine, OVA-G-L-serin: - 1,2-1,4 for 1:2500 Dilution, - 0,7-0,8 for 1:5000 Dilution - 0,4-0,5 for 1:10000 Dilution So the antibodies did not show an chiral discrimination between D-serine and L-serine coupled to BSA or OVA via glutaraldehyd. This results are in contradiction to your specification given in the product data sheet for ab6472 in which a clear discrimination between D-serine-G-BSA and L-serine-G-BSA is announced. I would be very pleased to if you advice me how the results have been acquired at abcom.

ANSWER: Thank you for enquiry, sorry to hear you are experiencing problems, we will do our best to resolve them. Please could you confirm whether the results listed below refer to repeats in the same dilution, i.e have you obtained exactly the same results for each Ab used at each dilution? Signals (Extinction)): For all haptens: BSA-G-D-serine, BSA-G-L-Serine, OVA-G-D-serine, OVA-G-L-serin: - 1,2-1,4 for 1:2500 Dilution, - 0,7-0,8 for 1:5000 Dilution - 0,4-0,5 for 1:10000 Dilution

ANSWER:

Thank you for your enquiry. Using conjugated D-Serine-Glutaraldehyde-Protein, antibody specificity was performed with an ELISA test by competition experiments with the following compounds:

Compound : Cross-reactivity ratio(a) D-Serine-G-BSA: 1 L-Serine-G-BSA: 1/>50000 D-Cysteine-G-BSA: 1/>50000 D-Alanine-G-BSA: 1/>50000

Below is an example of an ELISA protocol used to test conjugated D-Serine. If you have any additional questions, please contact us again.

1-Coating of conjugated D-Serine (10µg/ml) in maxisorp well plates (Nunc) with a solution of sodium carbonate buffer 0.05M (pH 9,6), during sixteen hours at 4°C. 2-Saturation of well plates with of a solution of PBS (pH 7,3) containing 1g/l of BSA (Acros), 10% of glycerol and 0.5% of Tween (one hour at 37°C). 3-Wash with PBS containing 0.5% of Tween (PBS Tween) (three times). 4-Preabsorbed D-Serine antiserum will be diluted (1/10,000-1/25,000) in PBS Tween containing 1g/l BSA, 1g/l of BSA-G and 10% of glycerol, 200µl by well plate (incubating during 2 hours at 37°C). 5-Wash with PBS Tween (three times). 6-200µl of peroxidase-labelled goat anti-rabbit (Jackson) diluted (1/10,000) in a solution of PBS Tween containing 1g/l of BSA, will be applied by well plate (during one hour at 37°C). 7-Well plates will be rinsed with PBS Tween (three times). 8-And finally the peroxidase will be developed by incubating 200µl by well plate of a citrate 0,1M/phosphate 0,2M (pH 5) solution containing 0.4% of OPD (Sigma) and 0.03% of hydrogen peroxide (Acros) for ten minutes in the dark, after that, we will stop the reaction by the addition of 50µl of 2M HCl. 9-The optical density will be mesured at 492nm.

Beginning of February 2004 I received D-serine Antibodies ab6472 which I would like to use to establish an D-serine ELISA for following D-serine blood level in patients with liver transplantation. System: Kompetitive ELISA Plate: Exiqon Coating: BSA-G-D-serine, BSA-G-L-Serine, OVA-G-D-serine, OVA-G-L-serine, over night, extensive washing with phoshate puffer Incubation of anti -D-serine antibodies (ab6472) for 3 hours, extensive washing with phoshate puffer, Tested dilution 1:2500, 1:5000, 1:10000 Incubation of anti-Rabbit-Antibodies conjugateted to HRP (used as Standard for other applications), 1 hour, extensive washing with phoshate puffer Addition of TMB (15 min.) and stopp-solution

Signals (Extinction)): For all haptens: BSA-G-D-serine, BSA-G-L-Serine, OVA-G-D-serine, OVA-G-L-serin: - 1,2-1,4 for 1:2500 Dilution, - 0,7-0,8 for 1:5000 Dilution - 0,4-0,5 for 1:10000 Dilution

So the antibodies did not show an chiral discrimination between D-serine and L-serine coupled to BSA or OVA via glutaraldehyd.

This results are in contradiction to your specification given in the product data sheet for ab6472 in which a clear discrimination between D-serine-G-BSA and L-serine-G-BSA is announced.

I would be very pleased to if you advice me how the results have been acquired at abcom.

ANSWER:

Thank you for enquiry, sorry to hear you are experiencing problems, we will do our best to resolve them. Please could you confirm whether the results listed below refer to repeats in the same dilution, i.e have you obtained exactly the same results for each Ab used at each dilution? Signals (Extinction)): For all haptens: BSA-G-D-serine, BSA-G-L-Serine, OVA-G-D-serine, OVA-G-L-serin: - 1,2-1,4 for 1:2500 Dilution, - 0,7-0,8 for 1:5000 Dilution - 0,4-0,5 for 1:10000 Dilution

We look forward to hearing from you.

My research need antibody agaist D-serine.I find the product in your company, But there isn't point the configuration of the amino acid.Now i need know wether the antibody is suitable to my research.In addition, can you send details about the procedure of the antibody against serine, such as the type of carrier protein and antigen dose every time immunization and so on.

ANSWER:

This antibody was raised against L-serine and only recognizes L-serine. We are in the process of raising antibodies against D-serine.