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TNFa _ TNF_alpha _ TNFSF2 ELISA Kit , Human ELISA kit
/Product Detail : SEKA10602 TNFa _ TNF_alpha _ TNFSF2 ELISA Kit , Human ELISA kit
Product name : TNFa _ TNF_alpha _ TNFSF2 ELISA Kit , Human ELISA kit
Catalog number : SEKA10602
Quantity: 5 Plates
Supplier name : Smart Serology
Data sheet : Ask more or other datasheet now !
TNFa _ TNF_alpha _ TNFSF2 ELISA Kit , Human ELISA kit Human samples 80 % of the research is conducted on human samples. Gentaur suppliers human normal cells, cell lines, RNA extracts and lots of antibodies and elisa kit to Human proteins as well as TNFa _ TNF_alpha _ TNFSF2 ELISA Kit , Human ELISA kit.
Human TNFa / TNF-alpha / TNFSF2
ELISA Pair Set
1. Materials provided
Capture Antibody - 0.4 mg/mL of mouse anti-TNFα monoclonal antibody. Dilute to a working concentration of 2.0 μg/mL in CBS before coating.
Detection Antibody - 0.4 mg/mL of mouse anti-TNFα monoclonal antibody conjugated to horseradish-peroxidase. Dilute to a working concentration of 1.0 μg/mL in detection antibody diluteion buffer before use
Standard - Each vial contains 34 ng of recombinant TNFα. Reconstitute with 1 mL detection antibody dilution buffer. After reconstitution, store at -20℃ to -70℃ in a manual defrost freezer. A six-point standard curve using 2-fold serial dilutions in sample dilution buffer, and a high standard of 500 pg/mL is recommended
The minimum detectable dose of human TNFa ( TNF-alpha / TNFSF2 ) was determined to be approximately 7.8125 pg/ml. This is defined as at least three times standard deviations above the mean optical density of 10 replicates of the zero standard
3. Principle of the product
The human TNFa ( TNF-alpha / TNFSF2 ) ELISA Pair Set is for the quantitative determination of human TNFa.
This ELISA Pair Set contains the basic components required for the development of sandwich ELISAs.
The Sino Biological ELISA Pair Set is a solid phase sandwich ELISA (Enzyme-Linked Immunosorbent Assay). It utilizes a monoclonal antibody specific for TNFa ( TNF-alpha / TNFSF2 ) coated on a 96-well plate. Standards and samples are added to the wells, and any TNFa present binds to the immobilized antibody. The wells are washed and a horseradish peroxidase conjugated mouse anti-TNFa monoclonal antibody is then added, producing an antibody-antigen-antibody "sandwich". The wells are again washed and TMB substrate solution is loaded, which produces color in proportion to the amount of TNFa present in the sample. To end the enzyme reaction, the stop solution is added and absorbances of the microwell are read at 450 nm
STORAGE - Detection Antibody should be protected from prolonged exposure to light. Aliquot the reagents and store at -20℃ to -70℃ in a manual defrost freezer.
1. Dilute the capture antibody to the working concentration in CBS. Immediately coat a 96-well microplate with 100μL per well of the diluted capture antibody. Seal the plate and incubate overnight at 4℃.
2. Aspirate each well and wash with at least 300μl wash buffer, repeating the process two times for a total of three washes. Complete removal of liquid at each step is essential for good performance. After the last wash, remove any remaining wash buffer by inverting the plate and blotting it against clean paper towels.
3. Block plates by adding 300 μL of blocking buffer to each well. Incubate at room temperature for a minimum of 1 hour.
4. Repeat the aspiration/wash as in step 2. The plates are now ready for sample addition.
1. Add 100 μL of sample or standards in sample dilution buffer per well. Seal the plate and incubate 2 hours at room temperature.
2. Repeat the aspiration/wash as in step 2 of plate preparation.
3. Add 100 μL of the detection antibody, diluted in antibody dilution buffer, to each well. Seal the plate and incubate 1 hour at room temperature.
4. Repeat the aspiration/wash as in step 2 of plate preparation.
5. Add 200 μL of substrate solution to each well. Incubate for 20 minutes at room temperature ( if substrate solution is not as requested, the incubation time should be optimized ). Avoid placing the plate in direct light.
6. Add 50 μL of stop solution to each well. Gently tap the plate to ensure thorough mixing.
7. Determine the optical density of each well immediately, using a microplate reader set to 450 nm.
Calculation of results
1. Calculate the mean absorbance for each set of duplicate standards, controls and samples. Subtract the mean zero standard absorbance from each.
2. Construct a standard curve by plotting the mean absorbance for each standard on the y-axis against the concentration on the x-axis and draw a best fit curve through the points on the graph.
3. To determine the concentration of the unknowns, find the unknowns’ mean absorbance value on the y-axis and draw a horizontal line to the standard curve. At the point of intersection, draw a vertical line to the x-axis and read the concentration. If samples have been diluted, the concentration read from the standard curve must be multiplied by the dilution factor.
4. Alternatively, computer-based curve-fitting statistical software may also be employed to calculate the concentration of the sample.
This standard curve is only for demonstration purposes. A standard curve should be generated for each assay.
Human TNFa / TNF-alpha / TNFSF2 ELISA Pair Set Flash
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