Schistosoma japonica polypeptide with immunogenicity and application thereof
An immunogenic and schistosome technology, applied in the field of bioengineering, can solve the problems of no research reports and achieve good immune prevention effect
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Embodiment 1
[0028] Example 1 Preparation of anti-Schistosoma japonicum adult soluble antigen antibody
[0029] 1. Preparation of Soluble Antigens from Schistosoma japonicum Adult Worms
[0030] 1) New Zealand white rabbits were infected with 2000 cercariae of Schistosoma japonicum by abdominal patch method, and were killed after 42 days. The worms were collected by portal vein perfusion and frozen in liquid nitrogen for later use.
[0031] 2) Take out two tubes of Schistosoma japonicum worm bodies from liquid nitrogen, and lyse the worm bodies with PBS (1×PBS, pH 7.0, 0.5Mm PMSF, 0.1mM EDTA).
[0032] 3) Use a glass homogenizer to grind the worm bodies, put them in an ice bath for 15 minutes of ultrasonic lysis, and repeat freezing and thawing at -70°C three times.
[0033] 4) Ultrasonic cracking in an ice bath for 15 minutes.
[0034] 5) Centrifuge at 12000g for 1 hour, and collect the supernatant as adult soluble antigen (SWA).
[0035] 6) Detect the OD260 and OD280 values of SWA w...
Embodiment 2
[0043] Example 2 Determination of titer of phage display cDNA library of adult Schistosoma japonicum (Sj44d)
[0044] The titer (titer) of phage is the number of live phage contained in one milliliter of culture medium. Phage counts can be performed because phages produce plaques visible to the naked eye on agar plates containing specific host bacteria. However, because the actual efficiency of the plaque counting method is difficult to approach 100% (generally low, because there are a few live phages that may not cause infection), in order to accurately express the concentration (titer or titer) of the virus suspension, it is generally not used The absolute number of virions is expressed in plaque-forming units (pfu for short). The specific steps of phage titer determination are as follows:
[0045] 1) Inoculate the host strain BLT5403 in 50ml of M9LB culture medium, shake at 37°C and 200rpm / min to make it OD 600nm The value reaches around 0.5;
[0046] 2) Take 10 μL of t...
Embodiment 3
[0050] Example 3 Screening of Schistosoma japonicum adult worm (Sj44d) phage display cDNA library
[0051] Based on the phage display cDNA library of Schistosoma japonicum adults (the phage display cDNA library of Schistosoma japonicum is preserved by our laboratory, and its library capacity is 4.98×10 6 pfu, the recombination rate is 93.8%), Schistosoma japonicum sclerotin was selected as a probe, and Schistosoma japonicum adult phage display cDNA library was screened in order to screen for receptor molecules related to S. The specific steps of method screening are as follows:
[0052] 1) Dilute the purified Schistosoma japonicum spore groove protein with TBS to a concentration of 1 μg / mL;
[0053] 3) Add 100 μL / well of Schistosoma japonicum dilute protein dilution to the ELISA plate, and coat overnight at 4°C;
[0054] 4) The next day, pour off the coating solution, pat dry, wash 5 times with TBST, shake on a shaker for 10 minutes each time;
[0055] 5) Add 150 μL of 3% B...
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