Background[1-3]
Rat creatine phosphate kinase (CPK) ELISA kit is used for in vitro quantitative detection of the content of rat creatine phosphate kinase (CPK) in serum, plasma, tissue homogenate and related liquid samples.
Experimental principle: The basis of ELISA is the solid phase of antigen or antibody and the enzyme labeling of antigen or antibody. The antigen or antibody bound to the surface of the solid phase carrier still retains its immunological activity, and the enzyme-labeled antigen or antibody retains both its immunological activity and the activity of the enzyme.
During the measurement, the specimen to be tested (the antibodies or antigens in it are measured) react with the antigens or antibodies on the surface of the solid chain extender carrier. The antigen-antibody complex formed on the solid-phase carrier is separated from other substances in the liquid by washing. Then add enzyme-labeled antigen or antibody, which is also bound to the solid-phase carrier through reaction.
At this time, the amount of enzyme on the solid phase is proportional to the amount of the test substance in the specimen. After adding the substrate for the enzyme reaction, the substrate is catalyzed by the enzyme into a colored product. The amount of the product is directly related to the amount of the test substance in the specimen, so qualitative or quantitative analysis can be performed based on the depth of the color.
Rat Creatine Phosphokinase (CPK) ELISA Kit
Operation steps
1. Adding standards: Set up standard wells and sample wells, and add 50 μL of standards of different concentrations to each standard well;
2. Add samples: Set up blank wells (no samples and enzyme-labeled reagents are added to the blank control wells, and the remaining steps are the same) and sample wells to be tested. First add 40 μl of sample diluent to the well of the sample to be tested on the enzyme-labeled coated plate, and then add 10 μl of the sample to be tested (the final dilution of the sample is 5 times). Add the sample to the bottom of the well of the enzyme plate, try not to touch the wall of the well, and shake gently to mix.
3. Add enzyme: Add 100 μl of enzyme label reagent to each well, except blank wells.
4. Incubation: Seal the plate with sealing film and incubate at 37°C for 60 minutes.
5. Solution preparation: Dilute the 20-fold concentrated washing solution with distilled water 20-fold and set aside.
6. Washing: Carefully remove the sealing film, discard the liquid, spin dry, fill each well with washing liquid, let it stand for 30 seconds and then discard, repeat this 5 times, and pat dry.
7. Color development: First add 50μl of chromogen A to each well, then add 50μl of chromogen B, shake gently to mix, and develop color for 15 minutes at 37°C in the dark.
8. Termination: Add 50 μl of stop solution to each well to terminate the reaction (blue turns to yellow immediately).
9. Measurement: Zero the blank wells and measure the absorbance (OD value) of each well at 450nm wavelength. The measurement should be performed within 15 minutes after adding the stop solution.
Apply[4][5]
Study on ventricular remodeling in rats with myocardial ischemia-reperfusion injury and the effects and mechanisms of mangiferin
Establishment and evaluation of the rat myocardial MIRI model. After ligating the LAD, a myocardial ischemia-reperfusion model was successfully established. Within 2-5 minutes after ligation, the electrocardiogram of the MIRI group showed an increase in QRS wave width, and the J point or ST segment elevation on the electrocardiogram was associated with The towering T-wave merges into a unidirectional curve upwards. Compared with the Sham group, the systolic and diastolic functions of the MIRI group decreased, as shown by the decrease in LVSP and ±dp/dtmax (P<0.05); and the increase in LVEDP (P<0.05).
The serum AST (871.43±69.54), CPK (1652.86±101.37) and LDH (2109.86±115.45) activities in the MIRI group were all increased compared with the Sham group (555.75±55.74, 1078.38±102.14, 1328.13±86.99) (P<0.05) ; The MDA content in myocardial tissue was increased compared with the Sham group (P<0.05), while the SOD and GSH-Px activities were decreased compared with the Sham group (P<0.05); the serum IL-6 and TXA2 contents were increased compared with the Sham group (P<0.05). 0.05), while the concentrations of IL-10 and PGI2 were lower than those in the Sham group (P<0.05);
Myocardial infarction area (MIS) in the MIRI group was (49.83±7.25%), and in the Sham group was 0%. There was a statistically significant difference between the two groups (P<0.05); in the MIRI group, the ultrastructural damage was severe and the sarcomere structure was arranged. Disorder, myofilaments dissolve, break, and disappear; mitochondria increase, swell, and cristae break; nuclear matrix cavitates, chromatin condenses, forming blocks and fragments of different sizes and shapes.
