AAV2 Titer Caspid ELISA Kit

Highlights

Wide Linear Range
Quantitation from 4.8×10⁷ to 2.5×10⁹ Capsids/mL

AAV2 Control Included
The AAV2 Control is provided together with kits

Great Assay Precision
With Intra-assay CV ≤ 10% and Inter-assay CV ≤ 15%

12 x 8-well Strips
Flexible format, only use the amount as it’s needed

Product List

Learn more about AAV2 Titer Caspid ELISA Kit

• Assay Principal

  AAV2 Titer Capsid ELISA Kit is based on the sandwich ELISA method.

  When standards (AAV2 Control) or samples are added to the capture plate, the AAV2 capsids in the sample can be captured on the plate coated with the capture antibody. Then the detection antibody (Biotin Anti-AAV2 Antibody) conjugated with biotin is added to interact with the AAV2 capsids bound on the plate. Streptavidin-Horseradish Peroxidase conjugate (Streptavidin-HRP) is added to interact with the biotin conjugated detection antibody. After washing steps, 3,3',5,5'-Tetramethylbenzidine solution (TMB Solution) is added resulting in formation of blue color. The reaction is stopped by adding Stop Solution. Application of the Stop Solution results in the color changing from blue to yellow. The intensity of the color can be read at 450 nm and 650 nm by a microplate reader. The quantity of AAV2 capsids in the sample is precisely quantified against an AAV2 standard curve.

  

  Figure 1: Schematic diagram of AAV2 Titer Capsid ELISA Kit

Validation Data

• Great Recovery Rate

  Recovery is a fundamental biophysical characteristic to evaluate immunoassay. Ideally, the recovery rate should be ultimately close to 100%. 100% recovery means there is no interference from the diluent or matrix in the assay. Normally, an 80% to 120% recovery rate can be regarded as there being relatively slight interference from the diluent or matrix. From the assay in triplicates, the recovery rates of GenScript AAV2 Titer Capsid ELISA kit are all located in the range of 80% to 120%, which indicates AAV2 titration with this ELISA kit is accurate and reproducible.

  Table 1: Recovery rate of AAV2 Titer Capsid ELSIA kit in triplicates

  Batch No.	Repeats	Average Conc. Capsids/mL	Recovery Rate
  			GenScript	Standard(80%-120%)
  #1	3	1.99E+09	93%	Qualified
  #2	3	2.05E+09	117%	Qualified
  #3	3	1.86E+09	95%	Qualified

• High Intra-assay and Inter-assay Precision

  A. High intra-assay precision

  The following are examples of reports and data generated from a representative intra-assay precision validation using AAV2 Titer Capsid ELISA kit for samples at low, medium, and high concentration levels. For testing, each sample was repeated 10 times, and the mean (M) and standard deviation (SD) of the 10 measured concentration results were calculated for each sample, and the coefficient of variation (CV) was calculated. Based on the evaluation results, the intra-assay % CVs are less than 10%, indicating good repeatability of AAV2 titration using the kit.

  Table 2: Intra-assay CV evaluation with three-batch AAV2 Titer Capsid ELISA kit

  Batch No.	Repeats	Average Conc. pg/mL	%CV
  			GenScript	Standard (≤10%)
  #1	10	1.37E+08	5%	Qualified
  #1	10	4.71E+08	3%	Qualified
  #1	10	1.87E+09	4%	Qualified
  #2	10	1.65E+08	3%	Qualified
  #2	10	5.15E+08	2%	Qualified
  #2	10	1.99E+09	3%	Qualified
  #3	10	1.41E+08	5%	Qualified
  #3	10	4.64E+08	4%	Qualified
  #3	10	1.79E+09	5%	Qualified

  B. High inter-assay precision

  The following are examples of reports and data generated from a representative inter-assay precision validation using 3 different batches of AAV2 Titer Capsid ELISA kit for the samples at low, medium, and high concentration. Each sample was tested for 10 replicates using the kit from three different batches, and the mean (M) and standard deviation (SD) of the 30 measured concentration results were calculated for each sample, and the coefficient of variation (CV) was calculated as well. Based on the results, the Inter-assay % CVs are less than 15%, indicating the kit has good inter-assay precision.

  Table 3: Inter-assay CV evaluation with three-batch AAV2 Titer Capsid ELISA kit

  Batch Amount	Repeats	Average Conc. pg/mL	%CV
  			GenScript	Standard (≤15%)
  3	3x10	1.47E+08	9%	Qualified
  3	3x10	4.83E+08	5%	Qualified
  3	3x10	1.88E+09	6%	Qualified

• Specificity/Cross-Reactivity

  The AAV2 Titration ELISA was performed according to the protocol stated in the product manual. As samples, empty AAV capsid preparations of serotypes 1, 2, 5, 6, 8, 9, and DJ with respective concentrations were applied to the microplate. The following ODs were obtained:

  Serotype	Capsids Titer	Absorbance (450/650 nm)
  	(Capsids/mL)	Duplicate 1	Duplicate 2	Average
  AAV 1	1.07E+10	0.0124	0.0118	0.0121
  AAV 2	1.20E+10	4.975	5.0119	4.99345
  AAV 5	4.29E+10	0.0149	0.0128	0.01385
  AAV 6	3.13E+10	0.0169	0.0153	0.0161
  AAV 8	2.31E+10	0.0162	0.013	0.0146
  AAV 9	1.22E+10	0.0138	0.014	0.0139
  AAV DJ	1.48E+10	0.0138	0.0131	0.01345
  Negative Control (Buffer Only)	0	0.0134	0.0114	0.0124

  The OD value for AAV2 capsids was above the negative control OD, whereas OD values for all other tested AAV serotypes were within the background range. The AAV2 Titer Capsid ELISA Kit is specifically calibrated for AAV2 titration, and cannot be applied for titer determination for other AAV serotypes.

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