Mouse IgE ELISA Kit

Sample collection, processing and preservation methods

1 Serum: use a test tube free of pyrogens and endotoxins, avoid any cell stimulation during operation, after collecting blood, 3000 Rotational centrifugation 10 Serum and red blood cells were rapidly and carefully separated.

2 Plasma: EDTA , citrate or heparin anticoagulation. 3000 Rotational centrifugation 30 The supernatant was removed in minutes.

3 Cell supernatant: 3000 Rotational centrifugation 10 Min to remove particles and polymers.

4 Tissue homogenization: add appropriate amount of normal saline to the tissue and mash it. 3000 Rotational centrifugation 10 The supernatant was removed in minutes.

5 Storage: If the sample is not tested in time after collection, please pack it according to one dosage and freeze it in -20℃, Avoid repeated freezing and thawing, thaw at room temperature and ensure the sample is thawed evenly and well.

Bring your own items

1. Microplate reader ( 450nm ）

2. High-precision sampler and gun head: 0.5-10uL 、 2-20uL 、 20-200uL 、 200-1000uL

3. 37℃ Incubator

Reagent Preparation:

20× Dilution of wash buffer: distilled water according to 1 ： 20 Dilution, i.e. 1 Portion of 20× washing Buffer Plus 19 Parts of distilled water.
Plate washing method:

1 Manual plate washing: throw out the liquid in the holes, fill each hole with washing liquid, and let it stand 1min Then throw out the liquid in the hole, pat it dry on absorbent paper, and wash the plate in this way 5 Times.

2 Automatic plate washing machine: inject washing solution into each hole 350μL , immersion 1min , wash plate 5 Times.

Operation steps:

1 Equilibration from room temperature 20min Take out the required slats from the aluminum foil bag after that, and seal the remaining slats with ziplock bag and put them back 4℃ 。

2 Place standard wells and sample wells, and add different concentrations of standards to each standard well 50 μ L ；

3 Add the sample to be tested first to the sample hole 10 μL , add sample diluent 40 μL ； Blank holes are not Plus.

4 Add horseradish peroxidase to each well of the standard well and sample well except for the blank well （ HRP ) Labeled detection antibody 100 μL Sealing the reaction hole with a sealing plate membrane, 37℃ Incubation in water bath or incubator 60min 。

5 Discard the liquid, pat dry on absorbent paper, fill each hole with washing liquid, and let stand 1min , throw away Wash liquid, pat dry on absorbent paper, and repeat washing plates 5 Times (the plate can also be washed with a plate washing machine).

6 Adding substrate per well A 、 B Each 50 μL ， 37℃ Incubate in the dark 15min 。

7 , adding stop solution to each well 50 μL ， 15min Inside, in 450nm Wavelength measurement of each well OD Value.

Result judgment

Draw the standard curve: in Excel In the worksheet, the standard concentration is used as the abscissa, corresponding to OD Use the value as ordinate, draw the linear regression curve of the standard product, and calculate the concentration value of each sample according to the curve equation.

Kit Performance

1 Accuracy: correlation coefficient between linear regression of standard and expected concentration R Value greater than or equal to 0.9900 。

2 Sensitivity: the lowest detection concentration is less than 0.1 μg/mL 。

3 Specificity: does not cross-react with other soluble structural analogs.

4 , repeatability: the intra-plate and inter-plate variation coefficients are less than 15% 。

5 Storage: 2-8℃ , Keep protected from light and moisture.

6 Expiry date: 6 Months