Monkey Major Histocompatibility Complex Class I (MHCI/HLA-I) ELISA Kit Instructions for Use

This kit can only be used for scientific research and should not be used for medical diagnosis.

Main Organization monkey (Monkey) MHC class I

(MHC I /HLA- I ) ELISA Test Kit

user's Guide

Detection principle

The kit uses a double antibody one-step sandwich enzyme-linked immunosorbent assay (ELISA). To the coated microwells pre-coated with the major histocompatibility complex class I (MHCI) antibody, the test specimens, standards, and HRP-labeled detection antibodies were sequentially added, incubated and thoroughly washed. Using the substrate TMB to develop color, TMB is converted to blue under the catalysis of peroxidase and converted to the final yellow color by the action of an acid. The color depth is positively correlated with the major histocompatibility complex class I (MHCI) in the sample. The absorbance (OD value) was measured at 450 nm using a microplate reader to calculate the sample concentration.

Sample collection, processing and storage methods

1. Serum: Use a tube containing no pyrogen and endotoxin. Avoid any cell irritation during the procedure. After collecting the blood, centrifuge and centrifuge for 10 minutes at 3000 rpm to quickly and carefully separate the serum and red blood cells.

2. Plasma: EDTA, citrate or heparin anticoagulation. The supernatant was taken by centrifugation at 3000 rpm for 30 minutes.

3. Cell supernatant: Centrifuge at 3000 rpm for 10 minutes to remove particles and polymer.

4. Tissue homogenization: The tissue is mashed by adding appropriate amount of physiological saline. The supernatant was taken by centrifugation at 3000 rpm for 10 minutes.

5. Storage: If the sample is not detected in time after collection, please dispense it once, freeze it at -20 °C, avoid repeated freezing and thawing, thaw at room temperature and ensure that the sample is fully thawed evenly.

Bring your own items

• Microplate reader (450nm)
• High-precision sampler and tip: 0.5-10uL, 2-20uL, 20-200uL, 200-1000uL
• 37 ° C incubator

Operational precautions

1. The kit was stored at 2-8 ° C and equilibrated for 20 minutes at room temperature before use. The concentrated washing liquid taken out from the refrigerator will crystallize, which is a normal phenomenon, and the water bath is heated to completely dissolve the crystals before use.
2. The slats not used in the experiment should be immediately put back into the ziplock bag and sealed (low temperature dry) for storage.
3. The S0 standard with a concentration of 0 can be regarded as a negative control or blank; the sample has been diluted 5 times according to the instructions, and the final result multiplied by 5 is the actual concentration of the sample.
4. Incubation is carried out in strict accordance with the time indicated in the instructions, the amount of liquid added and the order.
5. Shake well all liquid components before use.

Kit composition

Note: The concentration of standard (S0-S5) is: 0, 15, 30, 60, 120, 240 ng/mL

Reagent preparation

Dilution of 20× Wash Buffer: Distilled water was diluted 1:20, ie 1 part of 20× Wash Buffer plus 19 parts of distilled water.

Washing method

1. Manually wash the plate: Drain the liquid in the hole, fill each hole with the washing liquid, let stand for 1 min, then drain the liquid in the hole, pat dry on the absorbent paper, and wash the plate 5 times.
2. Automatic washing machine: Inject 350 μL of washing solution into each well, soak for 1 min, and wash the plate 5 times.

Steps

1. The required slats were taken out from the aluminum foil pouch after equilibrating for 20 min at room temperature, and the remaining slats were sealed back to 4 ° C with a ziplock bag.
2. Set standard and sample wells, standard wells with different concentrations of standard 50μL;
3. The sample well was first added with 10 μL of the sample to be tested, and then the sample dilution was 40 μL; the blank well was not added.
4. In addition to the blank wells, 100 μL of horseradish peroxidase (HRP)-labeled detection antibody was added to each well of the standard well and the sample well, and the reaction well was sealed with a sealing plate, and incubated at 37 ° C in a water bath or incubator for 60 min.
5. Discard the liquid, pat dry on the absorbent paper, fill each well with the washing solution, let stand for 1 min, remove the washing solution, pat dry on the absorbent paper, and repeat the washing 5 times (can also be washed with a washing machine).
6. 50 μL of each of the substrates A and B was added to each well, and incubated at 37 ° C for 15 min in the dark.
7. 50 μL of the stop solution was added to each well, and the OD value of each well was measured at a wavelength of 450 nm within 15 min.

Result judgment

Draw a standard curve: In the Excel worksheet, the standard concentration is used as the abscissa, and the OD value is plotted as the ordinate. The linear regression curve of the standard is drawn, and the concentration values ​​of each sample are calculated according to the curve equation.

Kit performance

• Accuracy: The linear regression coefficient of the standard and the expected concentration correlation coefficient R value, greater than or equal to 0.9900.
• Sensitivity: The minimum detection concentration is less than 1.0 ng/mL.
• Specificity: Does not cross-react with other soluble structural analogs.
• Repeatability: The coefficient of variation between the plates and the plates is less than 15%.
• Storage: 2-8 ° C, protected from light and moisture.
• Validity: 6 months

Disclaimer

• The kit is for research use only and should not be used for clinical trials or human experiments. Otherwise, the consequences will be borne by the experimenter and the company will not be responsible.
• In strict accordance with the instructions, the experimenter violates the instructions, and the consequences are borne by the experimenter.

Hematology Analyzer

Hematology analyzer is also called clinical blood cell analyzer, blood cell analyzer, blood cell analyzer, blood cell counter. The blood analyzer not only improves the accuracy of the experimental results, but also provides many experimental indicators, which play an important role in the diagnosis and differential diagnosis of diseases. Hematology analyzer is one of the most widely used instruments in hospital clinical testing.
Test items
Blood cell test refers to routine blood test, which is manual operation and counting under the microscope at first. It includes red blood cell, hemoglobin, white blood cell count and its classification, platelet count, etc. There are more than 20 items.
clinical significance
1. The blood analyzer is mainly used to detect various blood cell counts, white blood cell classification and hemoglobin content.
2. Hematocrit: obtained by multiplying the average volume of red blood cells by the red blood cell count.
3. Red blood cell distribution width: represents the degree of consistency of red blood cell size. When the red blood cell size is uneven, the red blood cell distribution width value increases, such as various types of nutritional deficiency anemia.
4. The three average indices of red blood cells are used to identify the type of anemia.
(1) The average hemoglobin content of red blood cells: increased in megaloblastic anemia, decreased in iron deficiency anemia, chronic blood loss anemia, uremia, chronic inflammation.
(2) Mean volume of red blood cells: increase in hemolytic anemia and megaloblastic anemia; decrease in severe iron deficiency anemia and hereditary spherocytosis.
(3) The average red blood cell hemoglobin concentration: decrease in chronic blood loss anemia, iron deficiency anemia; various diseases can be in the normal range. In megaloblastic anemia, the mean red blood cell volume increases, the mean red blood cell hemoglobin amount increases, the mean red blood cell hemoglobin concentration is normal, and the red blood cell distribution width increases.
5. Average platelet volume: the average volume of each platelet, the size of platelets is related to its function.
(1) Increased: seen in patients with idiopathic thrombocytopenic purpura, edema and proteinuria in late pregnancy.
(2) Decreased: seen in non-immune platelet destruction, aplastic anemia, thrombocytopenia repeated infection syndrome, chronic myeloid leukemia, etc.

Automation in Hematology,Automated Hematology Analyzer,Hematology Analyzer Instrument,Hematology Analyzer Product

Jilin Sinoscience Technology Co. LTD , https://www.contoryinstruments.com