Visible protein chip for detecting poultry disease serum antibody, its preparation method and application
A protein chip and protein technology, applied in the field of protein chips, can solve the problems of fast, accurate and high-throughput that cannot adapt to animal quarantine, and achieve the effects of good specificity, simplified preparation process and high reliability.
Inactive Publication Date: 2012-12-19
HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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Problems solved by technology
[0002] With the increasing frequency and scale of animal diseases in the world today, the classic serology and PCR detection methods can no longer meet the requirements of fast, accurate and high-throughput animal quarantine
Method used
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Embodiment 1
[0036] Embodiment 1 Preparation of protein chip of the present invention
[0037] Printing Buffer containing 1% BSA (137mM NaCl, 2.7mM KCl, 10mM NaCl 2 HPO 4 , 2mMKH 2 PO 4 , 1% BSA, 0.05% TW20) respectively diluted the purified virus antigen protein to 2000 μg / ml, and added it to a 384-well plate (20 μl per well). Keep the temperature at 25°C and the humidity at 50% to 60% to spot the sample on the aldylated glass slide, use SPF chicken serum as a positive control, Printing Buffer as a negative control, and repeat each virus antigen protein 6 times to form 6 ×6 protein chip array ( figure 1 ). After the samples were placed at room temperature for 1 hour, they were fixed in a drying box at 4°C for 24-48 hours. Use 1% BSA solution as the blocking agent, incubate in a 37°C incubator for 1 hour, take it out and wash it with PBS 3 times, 30 seconds each time.
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The invention discloses a visual protein chip for detecting serum antibody of new-castle disease virus of chickens, infectious bronchitis virus of chickens, avian influenza virus and infectious bursal disease virus of chickens , which is prepared by the following steps: purifying and diluting whole proteins of the four virus respectively; pointing samples of the positive control serum, the negative control serum and the four virus proteins onto a chip carrier respectively; drying, fixing, sealing and washing the samples to obtain the visual protein chip. The visual protein chip uses the purified whole proteins as capturing antigens to detect the virus-specific antibodies in chicken serum so as to simplify the preparation technology and reduce the production cost, and the visual protein chip has better specificity but no cross, has high reliability of results and has the advantages of quickness, simplicity and convenience, high sensitivity, good specificity and the like. When the serum is diluted by 6,400 times, the visual protein chip still can detect the antibodies, the sensitivity is 400 times of that of the prior AGP detection method. According to the detection to serum samples in-place, the detection rate of the visual protein chip is higher than the proir AGP method remarkably.
Description
technical field [0001] The present invention relates to a protein chip, in particular to a visualized protein chip for detecting 4 kinds of poultry disease serum antibodies. The invention also relates to a preparation method of the visualized protein chip and the application of the visualized protein chip in detecting 4 kinds of poultry disease serum antibodies, belonging to protein chip field. Background technique [0002] With the increasing frequency and scale of animal diseases in the world today, the classic serology and PCR detection methods can no longer meet the requirements of fast, accurate and high-throughput animal quarantine. Protein chip technology can meet the needs of simultaneous multi-index analysis of biological samples. It not only retains the rapidity, sensitivity and specificity of the antigen-antibody reaction, but also has the high throughput, low cost, high parallelism, miniaturization and extremely high performance of chip technology. Sensitivity a...
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IPC IPC(8): G01N33/531G01N33/569
Inventor 王秀荣包红梅石林伍嘉男陈化兰
Owner HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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