Trastuzumab (Herceptin) Immunogenicity ELISA Kit
适用物种:Human,mouse,rat

样本类型:Serum,Plasma

检测范围:125ng/ml - 31.25ng/ml

最低检出限:31.25ng/ml

测定时间:3.5 hours

检测格式:Bridging ELISA

检测原理:
The Trastuzumab immunogenicity assay employs the bridging ELISA technique. A precoated 96 well capture antibody plate is provided. Quality control and test samples are pipetted into the appropriate wells. Anti-Trastuzumab present in biological matrices binds the immobilized capture antibody. After washing away any unbound substances, secondary antibody is added to the wells and after a final wash a detection reagent is added. The plate is washed to remove any unbound antibody-enzyme reagent and a substrate solution is added to the wells for color development. The color development is proportional to the amount of anti-Trastuzumab present in test samples. Four levels of QC samples give a qualitative reference signal which can be used to determine the level (High, Medium, Low, Negative) of anti-Trastuzumab antibody in the unknown samples.

检测方法:Peroxidase / OD450 

靶抗原:Trastuzumab (Herceptin) Immunogenicity

应用:Quantification of antibodies to Trastuzumab

包被板类型:Strip

样品体积:15ul

特异性:Anti-Trastuzumab antibodies

检测精度(批内差异,批间差异):<10%, <10%

防腐剂:None

Gene ID:2064

试剂盒组分:
Coated microtiter plate, 96 wells
QC samples - 4x50ul
10X wash buffer - 25ml
Assay buffer - 50ml
1000X secondary antibody - 17ul
1000X detection reagent - 17ul
TMB - 12ml
TMB stop solution - 12ml
Plate sealers - 3

储存温度和稳定性:Stable at -20°C for 1 year

背景描述:
Trastuzumab (Herceptin® ) is a humanized recombinant monoclonal antibody used for the treatment of primary breast cancers overexpressing human epidermal growth factor 2 (HER2). HER2 protein is overexpressed in 25-30% of breast cancers.

检测程序:
This immunogenicity assay employs the bridging ELISA technique. Capture antibody is precoated onto a 96 well microplate. Quality control and test samples are pipetted into the appropriate wells. AntiTrastuzumab present in biological matrices is bound by the immobilized capture antibody. After washing away any unbound substances, secondary antibody is added to the wells and after a final wash a detection reagent is added. The plate is washed to remove any unbound antibody-enzyme reagent and a substrate solution is added to the wells for color development. The color development is proportional to the amount of anti-Trastuzumab present in test samples. Three levels of QC samples give a qualitative reference signal which can be used to determine the level of anti-Trastuzumab antibody in the unknown samples. The color development is stopped and the intensity of the color is measured.

样本收集:
This kit is compatible with EDTA-plasma, heparinplasma and serum samples. Samples can be stored at or below -20°C for up to 1 year.

样品制备:
Dilute QC samples and test samples 1/10 with assay buffer (for example add 30µL of prepared calibrator or sample to 270µL of assay buffer). Mix well. Do not store diluted samples. If test samples are out of range, then they may be further diluted.

试剂制备:
Prepare only the appropriate amount of required reagent on the day of use. Store all reagents as per instructions stated on the label.
1. Wash Buffer (1X) Preparation: Dilute wash buffer concentrate with ultra-pure water 1/10 before use (for example add 2mL concentrate to 18mL ultra-pure water). Mix well.
2. Secondary antibody (1X) Preparation: Dilute secondary antibody with assay buffer 1/1000 before use (for examples add 12μl concentrate to 12ml of assay buffer). Mix well.
3. Detection Reagent (1X) Preparation: Dilute detection reagent with assay buffer 1/1000 before use (for example add 12μl concentrate to 12ml of assay buffer). Mix well.

结果计算:
1. Because anti-drug antibodies will vary in terms of affinity and concentration, this assay provides a qualitative readout. As such the user should use the comparable positive controls when comparing interassay results. The provided controls are tested for comparability between lots and can be traced.
2. The anti-drug antibody titers in the test samples will fall in the range of high, medium, low or negative. We recommend each lab develop their own statistical cutpoint using methodologies as described by G. Shankar, et al. (2008). (Recommendations for the validation of immunoassays used for detection of host antibodies against biotechnology products. J. Pharmaceutical and Biomedical Analysis 48:1267–1281).
3. Any sample undiluted or diluted still reading greater than the highest standard should be diluted appropriately with assay buffer and retested. If the samples have been diluted, the concentration determined from the standard curve must be multiplied by the dilution factor.



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