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Mouse Immunoglobulin G2a (IgG2a) ELISA Kit


Species Reactivity

Mouse (Mus musculus)

UniProt

Abbreviation

IgG2a

Alternative Names

N/A

Range

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Sensitivity

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Sample Type

Serum, Plasma, Other biological fluids

Detection Method

Sandwich

Analysis Method

Quantitive

Assay Duration

1-4.5h

Sample Volume

1-200 μL

Detection Wavelengt

450 nm

Cat

AE64525MO

Size

96T/48T

Price

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Availability

3-5 working days

Manual

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Test principle:
This assay employs a two-site sandwich ELISA to quantitate IgG2a in samples. An antibody specific for IgG2a has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any IgG2a present is bound by the immobilized antibody. After removing any unbound substances, a biotin-conjugated antibody specific for IgG2a is added to the wells. After washing, Streptavidin conjugated Horseradish Peroxidase (HRP) is added to the wells. Following a wash to remove any unbound avidin-enzyme reagent, a substrate solution is added to the wells and color develops in proportion to the amount of IgG2a bound in the initial step. The color development is stopped and the intensity of the color is measured.
Product Overview:
BALB/c mice typically respond to vaccines and subunit vaccines with a Th2-type immune response, which is associated with the stimulation of IgG1 antibodies . However, the major antibody isotype present in the sera of mice that survive viral infections is IgG2a , which is stimulated during Th1-type immune responses . Stimulation of IgG2a antibodies has been associated with increased efficacy of vaccination . Additionally, monoclonal antibodies of the IgG2a isotype are more efficient at clearingvirus infections than monoclonal antibodies of the IgG1 isotype displaying similar antigenic specificities.
Aspace filling model of mouse IgG2a from the strain BALB/c. The two heavy chains are shown in yellow and light blue whilst the two light chains are shown in green and dark blue. The carbohydrate (red and green) is just visible in the Fc.

Components:

Reagents

Quantity

Reagents

Quantity

Assay plate (96 Wells)

1

Instruction manual

1

Standard (lyophilized)

2

Sample Diluent

1 x 20 mL

Biotin-Conjugate (concentrate 100 x)

1 x 120 μL

Biotin-Conjugate Diluent

1 x 20 mL

Streptavidin-HRP  (concentrate 100 x)

1 x 120 μL

Streptavidin-HRP Diluent   

1 x 20 mL

Wash Buffer (concentrate 25 x)

1 x 20 mL

Substrate Solution

1 x 12 mL

Stop Solution

1 x 10 mL

Adhesive Films

4

Specificity:
This assay has high sensitivity and excellent specificity for detection of Mouse IgG2a. No significant cross-reactivity or interference between Mouse IgG2a and analogues was observed.
Recovery:
Matrices listed below were spiked with certain level of recombinant Mouse IgG2a and the recovery rates were calculated by comparing the measured value to the expected amount of Mouse IgG2a in samples.

Sample Type

Number

Recovery range (%)

Average(%)

Serum

10

90-101

96

EDTA plasma

10

89-97

93

Heparin plasma

10

91-99

95

Precision:
Intra-assay Precision (Precision within an assay) Three samples of known concentration were tested twenty times on one plate to assess intra-assay precision. Inter-assay Precision (Precision between assays) Three samples of known concentration were tested in forty separate assays to assess inter-assay precision. CV (%) = SD/meanX100 Intra-Assay: CV<8% Inter-Assay: CV<12%
Linearity:
The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Mouse IgG2a and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.

Sample Type

1:2

1:4

1:8

1:16

Serum

78-89%

81-99%

92-103%

95-105%

EDTA plasma

91-101%

90-98%

93-101%

91-98%

Heparin plasma

92-103%

93-102%

92-99%

91-101%

Stability:
The stability of ELISA kit is determined by the loss rate of activity. The loss rate of this kit is less than 5% within the expiration date under appropriate storage condition. The loss rate was determined by accelerated thermal degradation test. Keep the kit at 37°C for 4 and 7 days, and compare O.D.values of the kit kept at 37°C with that of at recommended temperature. (referring from China Biological Products Standard, which was calculated by the Arrhenius equation. For ELISA kit, 4 days storage at 37°C can be considered as 6 months at 2 - 8°C, which means 7 days at 37°C equaling 12 months at 2 - 8°C).
Sample collection and storage:
Serum: Use a serum separator tube (SST) and allow samples to clot for two hours at room temperature or overnight at 2 - 8°C before centrifugation for 15 minutes at 1000 × g. Remove serum and assay immediately or aliquot and store samples at ≤ -20°C. Avoid repeated freeze-thaw cycles. Plasma: Collect plasma using EDTA, or heparin as an anticoagulant. Centrifuge for 15 minutes at 1000 × g at 2 - 8°C within 30 minutes of collection. Assay immediately or aliquot and store samples at ≤ -20°C. Avoid repeated freeze-thaw cycles. Other biological fluids: Centrifuge samples for 20 minutes at 1000 × g. Remove particulates and assay immediately or store samples in aliquot at -20°C or -80°C. Avoid repeated freeze/thaw cycles.
Kits storage instructions:
Store at 2-8°C. Please refer to Instruction Manual.