Monoclonal antibody hybridoma cell 3B5 strain capable of secreting H protein for resisting canine distemper virus

Abstract

The invention discloses a monoclonal antibody hybridoma cell 3B5 strain capable of secreting an H protein for resisting a canine distemper virus, and belongs to the biotechnology field. One hybridomacell strain 3B5 with a high neutralization activity is screened from an established bank of 127 strains of hybridoma cells, and an epitope is amino acids on the 228th-237th site of the H protein for resisting the canine distemper virus. The neutralization valence of an antibody composite therapeutic agent formed by the monoclonal antibody 3B5 and monoclonal antibodies 1D7 and 2G3 is 213, and a virus neutralization capability is improved by 8-32 times than the neutralization valence obtained when 3B5, 1D7 and 2G3 are independently used, the clinic application test effect of a monoclonal antibody composite therapeutic agent is improved to 100% from 98% than an effect obtained when the monoclonal antibodies 1D7 and 2G3 are subjected to combined utilization, and a treatment period is shortened to 2 days from 4 days. The monoclonal antibody hybridoma cell 3B5 strain can be used for the clinic treatment of CD (canine distemper) morbidity animals. By use of the strain disclosed by the invention, production cost is lowered, and a treatment effect is improved.

Description

technical field

[0001] The invention discloses a monoclonal antibody hybridoma cell 3B5 strain secreting anti-canine distemper virus H protein, and belongs to the field of biotechnology. Background technique

[0002] Canine distemper is an acute, febrile and highly contagious infectious disease caused by canine distemper virus (CDV), which often causes symptoms such as abnormalities in the digestive tract, respiratory tract and nervous system, and can cause immune suppression. The natural infection hosts of canine distemper have been expanded from the traditional Canidae, Weaselidae and Procyonidae to all carnivores, Artiodactyla Suidae, Primates Sages and Pinnipeds Sealidae and other animals, the incidence rate is almost It can reach 100%, and the mortality rate is more than 80%. It has become one of the important infectious diseases that endanger the dog industry and the economic animal breeding industry.

[0003] CDV belongs to the Paramyxoviridae family and the genus M...

Examples

Embodiment Construction

[0027] (1) Establishment of monoclonal antibody hybridoma cell lines

[0028] 1. Preparation of CDV antigen

[0029]Inoculate the CDV Onderstepoort strain (product of Holland Intervet International Co., Ltd.) into Vero cells (purchased from ATCC cell bank), collect the infection supernatant after the cells appear completely lesioned, freeze and thaw three times, and centrifuge at 8000r / min for 3 minutes. Remove cell debris; collect supernatant and add PEG 6000 (product of Sigma Company) to a final concentration of 9%, then sodium chloride was added to a final concentration of 3%, stirred at room temperature until dissolved, and precipitated overnight at 4°C. The next day, centrifuge at 8000r / min for 1 hour, discard the supernatant, and resuspend with 5mL TNE solution (pH7.8). Add the resuspension to the 60%-40%-20% sucrose gradient solution, 60000r / min ultracentrifugation for 3 hours, collect the layered protein bands (between 60%-40%), draw the virus bands, negative The vi...

technical field

[0001] The invention discloses a monoclonal antibody hybridoma cell 3B5 strain secreting anti-canine distemper virus H protein, and belongs to the field of biotechnology. Background technique

[0002] Canine distemper is an acute, febrile and highly contagious infectious disease caused by canine distemper virus (CDV), which often causes symptoms such as abnormalities in the digestive tract, respiratory tract and nervous system, and can cause immune suppression. The natural infection hosts of canine distemper have been expanded from the traditional Canidae, Weaselidae and Procyonidae to all carnivores, Artiodactyla Suidae, Primates Sages and Pinnipeds Sealidae and other animals, the incidence rate is almost It can reach 100%, and the mortality rate is more than 80%. It has become one of the important infectious diseases that endanger the dog industry and the economic animal breeding industry.

[0003] CDV belongs to the Paramyxoviridae family and the genus M...

Examples

Embodiment Construction

[0027] (1) Establishment of monoclonal antibody hybridoma cell lines

[0028] 1. Preparation of CDV antigen

[0029]Inoculate the CDV Onderstepoort strain (product of Holland Intervet International Co., Ltd.) into Vero cells (purchased from ATCC cell bank), collect the infection supernatant after the cells appear completely lesioned, freeze and thaw three times, and centrifuge at 8000r / min for 3 minutes. Remove cell debris; collect supernatant and add PEG 6000 (product of Sigma Company) to a final concentration of 9%, then sodium chloride was added to a final concentration of 3%, stirred at room temperature until dissolved, and precipitated overnight at 4°C. The next day, centrifuge at 8000r / min for 1 hour, discard the supernatant, and resuspend with 5mL TNE solution (pH7.8). Add the resuspension to the 60%-40%-20% sucrose gradient solution, 60000r / min ultracentrifugation for 3 hours, collect the layered protein bands (between 60%-40%), draw the virus bands, negative The vi...

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