CN104987418A - Method for producing anti-human retinol conjugated protein antiserum for sheep - Google Patents
Method for producing anti-human retinol conjugated protein antiserum for sheep Download PDFInfo
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Abstract
The invention provides a method for producing anti-human retinol conjugated protein antiserum for sheep. The method comprises the following steps of: (1) preparing an antigen; (2) performing injection for the first time; (3) performing injection for the second time; (4) performing injection for the third time; (5) performing injection for the fourth time; (6) performing injection for the fifth time; (7) performing injection for the sixth time; (8) performing injection for the seventh time; (9) performing injection for the eighth time; and (10) detecting serum titer: drawing blood from the vein of the sheep after 8-10 days from the injection each time, separating the serum and detecting dual-extending titer. Compared with the prior art, the method for producing the anti-human retinol conjugated protein antiserum for the sheep, disclosed by the invention, is low in cost, and simple in process; the method not only can stably produce the antibodies, but also is high in antibody producing speed; the antibody level is high, and the dosage of the antigen is small.
Description
Technical field
The present invention relates to a kind of sero-fast method of production goat-anti human retinol-binding protein, belong to antiserum prepare technical field.
Background technology
Retinol conjugated protein (Retinol-Binding Protein, RBP) is the translocator of VITAMIN in blood, is synthesized by liver, is distributed widely in blood, cerebrospinal fluid, urine and other body fluid.Measure the functional lesion of retinol conjugated protein energy early discovery uriniferous tubules, and the degree of damage of the sensitive reflection kidney proximal tubule of energy, also can be used as the index of liver function Random early Detection and monitoring and therapeutic.Its molecular weight is 21KD.
At present, after people RBP antigen and adjuvant emulsion, by certain program immunity animal, anti-human RBP antibody can be produced in animal serum.Although current production method can produce a certain amount of antibody, it is slow that antibody produces speed, and level is lower, and antigen consumption is high.
Summary of the invention
Goal of the invention: in order to overcome the deficiencies in the prior art, the invention provides a kind of sero-fast method of production goat-anti human retinol-binding protein.
Technical scheme: for achieving the above object, the invention provides a kind of sero-fast method of production goat-anti human retinol-binding protein, comprises the following steps:
(1) antigen prepares: select human retinol-binding protein antigen, carries out emulsification obtain antigen emulsion before injection with freund's adjuvant;
(2) first time injection: use above-mentioned antigen emulsion to inject sheep, injection site is vola, inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 10-16 point;
(3) second time injection: first time injects and carries out second time injection for latter 20 days, injection site is that lymphoglandula is interior, inguinal region, armpit, neck, dorsal sc multi-point injection, and each position 1-2 point, amounts to 10-16 point;
(4) third time injection: carry out third time injection after second time injection for 9-11 days, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 8-16 point;
(5) the 4th injections: carry out the 4th injection after third time injection for 9-11 days, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 8-16 point;
(6) the 5th injections: carry out the 5th injection after the 4th injection for 9-11 days, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 8-16 point;
(7) the 6th injections: carry out the 6th injection after the 5th injection for 9-11 days, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 8-16 point;
(8) the 7th injections: carry out the 7th injection after the 6th injection for 9-11 days, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 8-16 point;
(9) the 8th injections: carry out the 8th injection after the 7th injection for 9-11 days, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 8-16 point;
(10) serum titer detects: blood sampling in 8-10 days after per injection, separation of serum, detects its pair of expansion and tire.
As preferably, before described first time injection, the freund's adjuvant of emulsification is complete Freund's adjuvant, and for the second time, the freund's adjuvant of emulsification is incomplete Freund's adjuvant for the third time, before the 4th time, the 5th time, the 6th time, the 7th time and the 8th injection.
Preferred as another kind, the dosage of described first time injection is that every sheep amounts to 0.5mg antigen, each injection amount evenly distribute.
Preferred as another kind, the dosage of described second time injection is that every sheep amounts to 1.0mg antigen, each injection amount evenly distribute.
Preferred as another kind, the dosage of described third time, the 4th time, the 5th time, the 6th time, the 7th time and the 8th time injection is every sheep and amounts to 2.0mg antigen, each injection amount evenly distribute.
Preferred as another kind, the method that described serum titer detects is two expansion immunizations.
Preferred as another kind, described sheep is goat or sheep.
In the present invention, Freund's complete adjuvant is containing the cell wall constituent of mycobacterium tuberculosis, can stimulate division and the they produce antibodies of primed lymphocyte, cause the enlargement of lymphoglandula, strengthen the antibody response to antigen.
In addition, the present invention, by specific antigen dosage, immunization time, injection site and frequency injection, carries out immunity to sheep, finally obtains higher antibody horizontal.Emphasis of the present invention is that the antigen dose of every sheep injection in beginning three immunity is multiplied, with enhancement antigen gradually to the stimulation of body.
Beneficial effect: relative to prior art, the present invention produces the sero-fast method of goat-anti human retinol-binding protein, and cost is low, and process is simple, can not only stably manufactured antibody, and it is fast to produce antibody speed, and antibody horizontal is high, and antigen consumption is little.
Embodiment
Below in conjunction with embodiment, the present invention is further described.
Embodiment 1
Select 5 adult healthy goats, carry out the following step:
(1) antigen prepares: select containing human retinol-binding protein antigen, carries out emulsification obtain antigen emulsion before injection with freund's adjuvant;
(2) first time injection: get 2.5mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml complete Freund's adjuvant again and make antigen emulsion, this antigen emulsion is used to inject sheep, injection site is vola, inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 10-16 point, every sheep injected dose is that 2ml contains 0.5mg antigen, each injection amount evenly distribute;
(3) second time injection: first time injects and carries out second time injection for latter 20 days, get 5.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, this antigen emulsion is used to inject sheep, injection site is that lymphoglandula is interior, inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 10-16 point, every sheep injected dose is that 2ml contains 1.0mg antigen, each injection amount evenly distribute;
(4) third time injection: second time is injected and carried out third time injection for latter 9 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(5) the 4th injections: third time injects and carries out the 4th injection for latter 9 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(6) the 5th injections: the 5th injection is carried out in the 4th injection for latter 9 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(7) the 6th injections: the 6th injection is carried out in the 5th injection for latter 9 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(8) the 7th injections: the 7th injection is carried out in the 6th injection for latter 9 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(9) the 8th injections: the 8th injection is carried out in the 7th injection for latter 9 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(10) serum titer detects: blood sampling in 8 days after per injection, separation of serum, detects its pair of expansion and tire.
Embodiment 2
Select 5 adult healthy sheep, carry out the following step:
(1) antigen prepares: select containing human retinol-binding protein antigen, carries out emulsification obtain antigen emulsion before injection with freund's adjuvant;
(2) first time injection: get 2.5mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml complete Freund's adjuvant again and make antigen emulsion, this antigen emulsion is used to inject sheep, injection site is vola, inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 10-16 point, every sheep injected dose is that 2ml contains 0.5mg antigen, each injection amount evenly distribute;
(3) second time injection: first time injects and carries out second time injection for latter 20 days, get 5.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, this antigen emulsion is used to inject sheep, injection site is that lymphoglandula is interior, inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 10-16 point, every sheep injected dose is that 2ml contains 1.0mg antigen, each injection amount evenly distribute;
(4) third time injection: second time is injected and carried out third time injection for latter 11 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(5) the 4th injections: third time injects and carries out the 4th injection for latter 11 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(6) the 5th injections: the 5th injection is carried out in the 4th injection for latter 11 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(7) the 6th injections: the 6th injection is carried out in the 5th injection for latter 11 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(8) the 7th injections: the 7th injection is carried out in the 6th injection for latter 11 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(9) the 8th injections: the 8th injection is carried out in the 7th injection for latter 11 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(10) serum titer detects: blood sampling in 9 days after per injection, separation of serum, detects its pair of expansion and tire.
Embodiment 3
Select 5 adult healthy goats, carry out the following step:
(1) antigen prepares: select containing human retinol-binding protein antigen, carries out emulsification obtain antigen emulsion before injection with freund's adjuvant;
(2) first time injection: get 2.5mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml complete Freund's adjuvant again and make antigen emulsion, this antigen emulsion is used to inject sheep, injection site is vola, inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 10-16 point, every sheep injected dose is that 2ml contains 0.5mg antigen, each injection amount evenly distribute;
(3) second time injection: first time injects and carries out second time injection for latter 20 days, get 5.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, this antigen emulsion is used to inject sheep, injection site is that lymphoglandula is interior, inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 10-16 point, every sheep injected dose is that 2ml contains 1.0mg antigen, each injection amount evenly distribute;
(4) third time injection: second time is injected and carried out third time injection for latter 10 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(5) the 4th injections: third time injects and carries out the 4th injection for latter 10 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(6) the 5th injections: the 5th injection is carried out in the 4th injection for latter 10 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(7) the 6th injections: the 6th injection is carried out in the 5th injection for latter 10 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(8) the 7th injections: the 7th injection is carried out in the 6th injection for latter 10 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(9) the 8th injections: the 8th injection is carried out in the 7th injection for latter 10 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(10) serum titer detects: blood sampling in 10 days after per injection, separation of serum, detects its pair of expansion and tire.
Embodiment 4 (control group)
Select 5 adult healthy goats, carry out the following step:
(1) antigen prepares: select containing human retinol-binding protein antigen, carries out emulsification obtain antigen emulsion before injection with freund's adjuvant;
(2) first time injection: get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml complete Freund's adjuvant again and make antigen emulsion, this antigen emulsion is used to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(3) second time injection: first time injects and carries out second time injection for latter 20 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(4) third time injection: second time is injected and carried out third time injection for latter 10 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(5) the 4th injections: third time injects and carries out the 4th injection for latter 10 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(6) the 5th injections: the 5th injection is carried out in the 4th injection for latter 10 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(7) the 6th injections: the 6th injection is carried out in the 5th injection for latter 10 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(8) the 7th injections: the 7th injection is carried out in the 6th injection for latter 10 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(9) the 8th injections: the 8th injection is carried out in the 7th injection for latter 10 days, get 10.0mg human retinol-binding protein antigen, with PBS or normal saline dilution to 5ml, add 5ml incomplete Freund's adjuvant again and make antigen emulsion, use this antigen emulsion to inject sheep, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amount to 8-16 point, every sheep injected dose is that 2ml contains 2.0mg antigen, each injection amount evenly distribute;
(10) serum titer detects: blood sampling in 9 days after per injection, separation of serum, detects its pair of expansion and tire.
The two immunization that expands of experimental example detects antiserum titre
Vein adopts sheep blood 1ml, and separation of serum is about 0.5ml; By sheep blood serum PBS or physiological saline doubling dilution, be not respectively 1:2,1:4 to 1:128 not etc.; Beat plum blossom hole on a lbmc agar plate, aperture 2mm, pitch-row 3mm; Be sequentially added into 1:2,1:4 to 1:128 sheep blood serum 20 μ l not etc. in the hole outer Saturday often organizing plum blossom hole respectively, add at medium pore the human retinol proteantigen 20 μ l that concentration is 1.0mg/ml; The agar plate having added serum and antigen being put into 37 DEG C and hatches 12-24 hour, there is the final position of precipitation line in observation antigen and serum, and two expansion theing be serum is tired.
Control group is according to the immunity of embodiment 4 method, i.e. the antigen dose of each immunity is every sheep 2.0mg, and per injection position is inguinal region, armpit, neck, dorsal sc multi-point injection, does not carry out injection in vola and lymphoglandula;
Embodiment 1 group, 2 groups and 3 groups carry out immunization according to the embodiment of the present invention 1,2 and 3 prescription method respectively.
Detect each group of sero-fast two expansion of final gained by above method to tire, the results are shown in Table 1.
Table 1 respectively sero-fast two expansion of group gained is tired
Can be obtained by upper table 1 result, the antiserum(antisera) of control group gained, only have one to reach 1:32 only, other be 1:16; And in the present invention's three embodiment groups, 1:32 and 1:64 can be reached, gained antibody horizontal improves further greatly on the basis of control group, and antigen amount minimizing 2.5mg (16%) that every sheep uses.
Claims (7)
1. produce the sero-fast method of goat-anti human retinol-binding protein, it is characterized in that, comprise the following steps:
(1) antigen prepares: select human retinol-binding protein antigen, carries out emulsification obtain antigen emulsion before injection with freund's adjuvant;
(2) first time injection: use above-mentioned antigen emulsion to inject sheep, injection site is vola, inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 10-16 point;
(3) second time injection: first time injects and carries out second time injection for latter 20 days, injection site is that lymphoglandula is interior, inguinal region, armpit, neck, dorsal sc multi-point injection, and each position 1-2 point, amounts to 10-16 point;
(4) third time injection: carry out third time injection after second time injection for 9-11 days, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 8-16 point;
(5) the 4th injections: carry out the 4th injection after third time injection for 9-11 days, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 8-16 point;
(6) the 5th injections: carry out the 5th injection after the 4th injection for 9-11 days, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 8-16 point;
(7) the 6th injections: carry out the 6th injection after the 5th injection for 9-11 days, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 8-16 point;
(8) the 7th injections: carry out the 7th injection after the 6th injection for 9-11 days, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 8-16 point;
(9) the 8th injections: carry out the 8th injection after the 7th injection for 9-11 days, injection site is inguinal region, armpit, neck, dorsal sc multi-point injection, each position 1-2 point, amounts to 8-16 point;
(10) serum titer detects: blood sampling in 8-10 days after per injection, separation of serum, detects its pair of expansion and tire.
2. the sero-fast method of production goat-anti human retinol-binding protein according to claim 1, it is characterized in that, before the injection of described first time, the freund's adjuvant of emulsification is complete Freund's adjuvant, and for the second time, the freund's adjuvant of emulsification is incomplete Freund's adjuvant for the third time, before the 4th time, the 5th time, the 6th time, the 7th time and the 8th injection.
3. the sero-fast method of production goat-anti human retinol-binding protein according to claim 1, is characterized in that, the dosage of described first time injection is that every sheep amounts to 0.5mg antigen, each injection amount evenly distribute.
4. the sero-fast method of production goat-anti human retinol-binding protein according to claim 1, is characterized in that, the dosage of described second time injection is that every sheep amounts to 1.0mg antigen, each injection amount evenly distribute.
5. the sero-fast method of production goat-anti human retinol-binding protein according to claim 1, it is characterized in that, the dosage of described third time, the 4th time, the 5th time, the 6th time, the 7th time and the 8th time injection is every sheep and amounts to 2.0mg antigen, each injection amount evenly distribute.
6. the sero-fast method of production goat-anti human retinol-binding protein according to claim 1, is characterized in that, the method that described serum titer detects is two expansion immunizations.
7. the sero-fast method of production goat-anti human retinol-binding protein according to claim 1, is characterized in that, described sheep is goat or sheep.
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| CN107163145A (en) * | 2017-05-27 | 2017-09-15 | 海奥斯生物科技镇江有限公司 | One kind production sero-fast method of goat-anti human serum amyloid A |
| CN107163141A (en) * | 2017-05-27 | 2017-09-15 | 海奥斯生物科技镇江有限公司 | A kind of purification process of sheep polyvalent antibody specific antibody |
| CN107216386A (en) * | 2017-05-27 | 2017-09-29 | 海奥斯生物科技镇江有限公司 | A kind of goat-anti human serum prepares the accurate determination method of antigen immune dosage |
| CN107312086A (en) * | 2017-05-27 | 2017-11-03 | 海奥斯生物科技镇江有限公司 | The preparation method for the animal anti-human serum being precisely controlled based on immune position point |
| CN107383191A (en) * | 2017-05-27 | 2017-11-24 | 海奥斯生物科技镇江有限公司 | Gamut sheep or the preparation method of rabbit-anti Human C-reactiveprotein antibody |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN107163140A (en) * | 2017-05-27 | 2017-09-15 | 海奥斯生物科技镇江有限公司 | A kind of sero-fast preparation method of goat-anti human muscle hemoglobin |
| CN107163145A (en) * | 2017-05-27 | 2017-09-15 | 海奥斯生物科技镇江有限公司 | One kind production sero-fast method of goat-anti human serum amyloid A |
| CN107163141A (en) * | 2017-05-27 | 2017-09-15 | 海奥斯生物科技镇江有限公司 | A kind of purification process of sheep polyvalent antibody specific antibody |
| CN107216386A (en) * | 2017-05-27 | 2017-09-29 | 海奥斯生物科技镇江有限公司 | A kind of goat-anti human serum prepares the accurate determination method of antigen immune dosage |
| CN107312086A (en) * | 2017-05-27 | 2017-11-03 | 海奥斯生物科技镇江有限公司 | The preparation method for the animal anti-human serum being precisely controlled based on immune position point |
| CN107383191A (en) * | 2017-05-27 | 2017-11-24 | 海奥斯生物科技镇江有限公司 | Gamut sheep or the preparation method of rabbit-anti Human C-reactiveprotein antibody |
| CN107216386B (en) * | 2017-05-27 | 2021-02-12 | 海奥斯生物科技镇江有限公司 | Method for accurately determining antigen immunization dose prepared from sheep anti-human serum |
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Application publication date: 20151021 |