Anti-IgE抗体[5D4] (ab7295)

Thank you for contacting us.   I have received the following information from the lab: We are using ab7295 according to the datasheet recommendation as the capture antibody, not as detection antibody. The detection antibody ab7295 was used with the HRP-conjugated antibody ab7294. With this antibody pair we get the standard curve that is attached to this email.    We have used very standard internal protocol without any optimization: capture antibody ab7295 at 5 ug/ml in PBS, human IgE from 100 ng/ml to 1.56 ng/ml, detection antibody ab7294 (HRP) at 0.7 ug/ml concentration.    Unfortunately we don’t have any results using ab7295 as detection antibody. I am sorry for this inconvenience. However, it seems that levels of IgE much lower than 5 ug/mL or even 0.37 ug/mL are detectable with ab7295 used as capture antibody. I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information. 

Thank you for your enquiry. In our internal assay for anti-IgE MAb testing, highly purified myelomatous IgE(VL) was adsorbed on solid phase from solution (5 microgram/ml) in bicarbonate buffer pH 9.5. Ab7295 was applied for 60-120 min onto antigen and after washing HRP labeled affinity purified rabbit anti-mouse Ig antibody in TRIS-Tween buffer was used as developing reagent. The value 1:1 000 000 means the end-point of titration in conditions mentioned above. Generally ab7295 is recommended as capture antibody. Its concentration for immobilization onto solid phase may be 5-10 microgram/ml depending on kind of plastic plates used by customer. I hope this information helps. Please do not hesitate to contact us again if you need anything further.

Thank you for your enquiry and for completing our technical questionnaire. I am sorry to hear that you have been having difficulties with this antibody. I would greatly appreciate it if you could tell me of the concentration that you have been using this antibody at. Our datasheet states a 1/ 1 x 10^6 dilution. However, I consider this exceptionally dilute even by its application using ELISA. Your details of the concentration that you have been using was unfortunately lost in the formating of the e-mail. Please can you therefore tell me whether you have been using this antibody to coat the wells at 1ng/ml or 1ug/ml. I look forward to hearing from you.