Rat Visceral adipose-specific serine protease inhibitor,vaspin ELISA Kit使用说明书本

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Rat Visceral adipose-specific serine protease inhibitor,vaspin ELISA Kit使用说明书本
规格：48T/96T
货号：YM-SQ1785
全称：大鼠内脏脂肪特异性丝氨酸蛋白酶YZ剂(vaspin)ELISA 试剂盒
Rat Visceral adipose-specific serine protease inhibitor,vaspin ELISA Kit使用说明书本 Kit composition:
Reagent box is composed of 48 holes with 96 well configuration.
Instruction manual 1 1
Plate film 2 (48) 2 (96)
Sealed bag 1 1
The enzyme labeled plate was kept at 1 * 481 * 96 2-8
Standard product: 0.5ml 540ng/ml * 1 0.5ml * 1 2-8
Standard solution 1.5ml * 1 1.5ml * 1 2-8
Enzyme labeled reagent 3 ml * 6 ml * 1 2-8
Sample dilution liquid 3 ml * 6 ml * 1 2-8
Color ml 3 ml 6 A 1 2-8 1
Color ml 3 ml 6 B 1 2-8 1
3l * 1 6ml * 1 2-8
Concentrated washing solution (20ml * 20) * 1 (20ml * 30) x 1 bottles of 2-8
Sample handling and requirements:
1 serum: 10-20 minutes at room temperature and blood natural coagulation, about 20 minutes (2000-3000). Carefully collected supernatant, preserved in the process, such as precipitation, should be re centrifuge.
2. The plasma should be according to the requirements of the specimens of choice of EDTA or sodium citrate as an anticoagulant, 10-20 minutes after mixing, centrifuged for 20 minutes or so (2000-3000 r.p.m.). Carefully collected supernatant, preserved in the process, such as precipitation, it should be re - centrifugal.
3 urine: use sterile tube collection, centrifugal 20 minutes or so (2000-3000). Carefully collected supernatant, preserved in the process, such as precipitation, should be re - centrifugal. Reference to the chest and ascites, cerebrospinal fluid.
4 cell culture supernatant: the collection of sterile tubes for the detection of secretory components. Centrifugal 20 minutes or so (2000-3000 / min).
Carefully collected supernatant. When the cells were detected by PBS (PH7.2-7.4), the cell concentration reached 1000000 /ml. By repeatedly freezing and thawing, so that the cell damage and release of intracellular components. Centrifugal 20 minutes or so (2000-3000 / min). Carefully collected supernatant. In the process of preservation, if there is a precipitate, it should be again.
5 tissue specimens: after cutting the specimen, the weight is weighed. Adding a certain amount of PH7.4, PBS. Quickly frozen with liquid nitrogen. The temperature of the sample was maintained at 2-8. Adding a certain amount of PBS (PH7.4), using the hand or the homogenate of the specimen homogenate.
Centrifugal 20 minutes or so (2000-3000 / min). Carefully collected supernatant. Be detected after a repackaging, remaining frozen spare.
6 samples collected as soon as possible after extraction, extraction by the relevant literature, and then the experiment should be carried out as soon as possible. If the test can not be carried out immediately, the specimen can be kept at -20, but it should be avoided.
7 could not detect the NaN3 containing samples, because NaN3 inhibited the content of horseradish peroxidase (HRP).Operation steps
标准品的稀释与加样：在酶标包被板上设标准品孔10孔，在DY、第二孔中分别加标准品100μl，然后在DY、第二孔中加标准品稀释液50μl，混匀；然后从DY孔、第二孔中各取100μl分别加到第三孔和第四孔，再在第三、第四孔分别加标准品稀释液50μl，混匀；然后在第三孔和第四孔中先各取50μl弃掉，再各取50μl分别加到第五、第六孔中，再在第五、第六孔中分别加标准品稀释液50ul，混匀；混匀后从第五、第六孔中各取50μl分别加到第七、第八孔中，再在第七、第八孔中分别加标准品稀释液50μl，混匀后从第七、第八孔中分别取50μl加到第九、第十孔中，再在第九第十孔分别加标准品稀释液50μl，混匀后从第九第十孔中各取50μl弃掉。 (the L, 240ng/ml,, 60ng/ml,, 30ng/ml,, 360ng/ml,,,, and 120ng/ml, respectively)
Add sample: respectively, the blank hole (blank control hole without the sample and the enzyme label, and the remaining steps are the same), the sample hole. In the enzyme standard coated plate to be tested on a sample hole Zhongxian sample dilution of 40 g l, and then to be measured is added 10 mu l of sample (sample final dilution degrees for 5 times). The sample will be added to the bottom of the plate hole, and the hole wall is not to be touched.
Temperature: 37 minutes after the closure of the sealing plate with a sealing plate.
With liquid: 30 (48T 20 times) times concentrated washing liquid with distilled water 20 (48T 30 times) after dilution.
Washing: be careful torn off the seal plate membrane, discard liquid, drying, washing liquid to fill each hole, standing for 30 seconds after the discard, repeat 5 times, pat dry.
Add enzyme: 50 L per hole, except for the blank.
Wen Yu: operation with 3.
Washing: operation with 5.
Color: each hole first add color agent A50 L, and then add color agent B50 L, gently shake mix well, 37 to 15 minutes.
Termination: termination of 50 L per hole, terminate the reaction (at this time, blue).
Determination: the blank absorbance at 450nm air conditioning zero, in order to measure the hole (OD). Determination should be carried out within 15 minutes after the termination of the liquid.
Note:
Kit from the cold storage environment in the room temperature balance 15-30 minutes after the use of the enzyme labeled package is not used up after the Kaifeng, the plate should be stored in a sealed bag.
Washing buffer will crystallization, heated the water solubilization when diluted, washing does not affect the results.
Each step should be used with the sample, and often to check the accuracy, to avoid the test error. A sample within 5 mins, ifthenumberofsampleismuch, recommend to use volley like.
Please do the standard curve at the same time, it is best to do the hole. Such as samples to be measured matter content is too high (the sample od is bigger than the first standard hole hole OD), please first sample dilution multiples (n times) were measured and calculated please then multiplied by the total dilution ratio (n * * 5).
Closure plate membrane for one-time use, to avoid cross contamination.
Substrate please avoid light preservation.
In strict accordance with the instructions of the operation, the test results must be determined by the reader's reading.
All samples, washing liquid and all kinds of waste should be treated according to the transmission.
This reagent not mix different batches.
10 if there is a different from the instruction of the English language, the English instruction manual shall prevail.
Calculation:

To the standard concentration as the abscissa, OD value as the ordinate, draw the standard curve on coordinate paper, according to the OD value of samples from the standard curve found corresponding concentration; multiply again
Kit performance:
1 sample linear regression with the expected concentration correlation coefficient R value is 0.92 above.
The 2 batch and batch shall be less than 9% and 15% respectively.
Detection range:
-480ng/ml 12ng/ml
Preservation conditions and validity:
1 Kit: 2-8.
2 validity: 6 months

大鼠内脏脂肪特异性丝氨酸蛋白酶YZ剂(vaspin)ELISA 试剂盒