Active Member
|
[China]
Address: #1008.Junjiang Internatioanl Bldg. 228 Ningguo Rd. Yangpu Dist. Shanghai. 200090. China
Contact name:
Shanghai Korain Biotech Co., Ltd |
|
|
96 Well Plate Human High Precision and Sensitivity 5-Nucleotidase
ELISA Assay Kit For Research
*This product is for research use only, not for use in diagnosis
procedures. It’s highly recommend to read this instruction entirely
before use.
Intended Use
This sandwich kit is for the accurate quantitative detection of
human 5-Nucleotidase (also known as 5-NT) in serum, plasma, cell
culture supernates, cell lysates, tissue homogenates.
Assay Principle
This ELISA Assay Kit is an Enzyme-Linked Immunosorbent Assay (ELISA). The plate has been
pre-coated with human 5-NT antibody. 5-NT present in the sample is
added and binds to antibodies coated on the wells. And then
biotinylated human 5-NT Antibody is added and binds to 5-NT in the
sample. Then Streptavidin-HRP is added and binds to the
Biotinylated 5-NT antibody. After incubation unbound
Streptavidin-HRP is washed away during a washing step. Substrate
solution is then added and color develops in proportion to the
amount of human 5-NT. The reaction is terminated by addition of
acidic stop solution and absorbance is measured at 450 nm.
Reagent Preparation
All reagents should be brought to room temperature before use.
Standard Reconstitute the 120μl of the standard (160ng/ml) with 120μl of
standard diluent to generate a 80ng/ml standard stock solution.
Allow the standard to sit for 15 mins with gentle agitation prior
to making dilutions. Prepare duplicate standard points by serially
diluting the standard stock solution (80ng/ml) 1:2 with standard
diluent to produce 40ng/ml, 20ng/ml, 10ng/ml and 5ng/ml solutions.
Standard diluent serves as the zero standard(0 ng/ml). Any
remaining solution should be frozen at -20°C and used within one
month. Dilution of standard solutions suggested are as follows:
| 80ng/ml | Standard No.5 | 120μl Original Standard + 120μl Standard Diluent |
| 40ng/ml | Standard No.4 | 120μl Standard No.5 + 120μl Standard Diluent |
| 20ng/ml | Standard No.3 | 120μl Standard No.4 + 120μl Standard Diluent |
| 10ng/ml | Standard No.2 | 120μl Standard No.3 + 120μl Standard Diluent |
| 5ng/ml | Standard No.1 | 120μl Standard No.2 + 120μl Standard Diluent |
| Standard Concentration | Standard No.5 | Standard No.4 | Standard No.3 | Standard No.2 | Standard No.1 |
| 160ng/ml | 80ng/ml | 40ng/ml | 20ng/ml | 10ng/ml | 5ng/ml |
Wash Buffer Dilute 20ml of Wash Buffer Concentrate 25x into deionized or
distilled water to yield 500 ml of 1x Wash Buffer. If crystals have
formed in the concentrate, mix gently until the crystals have
completely dissolved.
Cat.No E0831Hu
Standard Curve Range: 0.5ng/ml - 150ng/ml
Sensitivity: 0.34ng/ml
Reagent Provided
| Components | Quantity |
| Standard Solution (160ng/ml) | 0.5ml x1 |
| Pre-coated ELISA Plate | 12 * 8 well strips x1 |
| Standard Diluent | 3ml x1 |
| Streptavidin-HRP | 6ml x1 |
| Stop Solution | 6ml x1 |
| Substrate Solution A | 6ml x1 |
| Substrate Solution B | 6ml x1 |
| Wash Buffer Concentrate (25x) | 20ml x1 |
| Biotinylated human 5-NT Antibody | 1ml x1 |
| User Instruction | 1 |
| Plate Sealer | 2 pics |
| Zipper bag | 1 pic |
Material Required But Not Supplied
Summary
1. Prepare all reagents, samples and standards.
2. Add sample and ELISA reagent into each well. Incubate for 1 hour
at 37°C.
3. Wash the plate 5 times.
4. Add substrate solution A and B. Incubate for 10 minutes at 37°C.
5. Add stop solution and color develops.
6. Read the OD value within 10 minutes.
Calculation of Result
Construct a standard curve by plotting the average OD for each
standard on the vertical (Y) axis against the concentration on the
horizontal (X) axis and draw a best fit curve through the points on
the graph. These calculations can be best performed with
computer-based curve-fitting software and the best fit line can be
determined by regression analysis.
