RU2380115C1 - Method of blood sampling for making antigens of haemosporidia - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: invention concerns biotechnology. The method involves splenectomy of an animal, infection with blood parasites, exsanguinations at invasion peak, blood conservation and use for making antigens. Thus before infection, autoblood is accumulated by fivefold draw and transfusion of autoblood in amount 150, 300, 500, 700 and 1000 ml respectively with isotonic solution every 3-4 days, then an animal is infected with Babesia canis. At parasitemia peak, when 30-50% of erythrocytes involved, the blood is taken twice every 3-4 days in amount 800-900 ml, and then the prepared autoblood is injected. Then at the third parasitemia peak, the animal is exsanguinated completely, and the blood prepared at three stages is used for making antigen.

EFFECT: method allows producing greater blood volume from one animal.

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Description

The present invention relates to the field of veterinary protozoology, in particular the production of blood for the manufacture of antigens from hemosporidia.

Dog babesiosis (pyroplasmosis) is a natural focal protozoal transmissible non-contagious blood-parasitic disease caused by the simplest parasite Babesia canis. The disease proceeds super-construction (lightning fast), acutely and chronically. The defeat of the body of the dog Babesia canis causes fever, a sharp increase in body temperature to 41-42 ° C, held for 2-3 days, followed by a rapid drop to and below normal. In some very rare cases, the temperature curve is less regular; the temperature undergoes large fluctuations, remains at a high level, after which there is a slow and gradual decrease. In very young dogs, in which death occurs very quickly, there may be no increase in temperature at the onset of the disease.

In dogs, there is a lack of appetite, depression, depression, a weak filiform pulse (up to 120-160 beats per minute), later it becomes arrhythmic. Heart beat boosted. Breathing is rapid (up to 36-48 per minute) and difficult, in young dogs often with a moan. On palpation of the left abdominal wall (behind the costal arch), an enlarged spleen is detected. The mucous membranes of the oral cavity and conjunctiva are anemic, icteric. Intensive destruction of red blood cells is accompanied by jade. The gait becomes difficult, hemoglobinuria appears. The disease lasts from 2 to 5 days, less often 10-11 days, often fatal.

According to many authors, this disease accounts for 14 to 25% of the total number of dogs that were provided with veterinary services. According to statistics over the past 10 years, the incidence of dogs with babesiosis has increased several times [3].

Serological reactions are widely used for the diagnosis of dogs babesiosis: the long-term complement binding reaction (RDSK), the indirect hemagglutination reaction (RIGA), enzyme-linked immunosorbent assay (ELISA), etc. The formulation of these reactions involves the use of Babesia canis antigens obtained from the blood of infected animals.

There is a method according to which a sick animal is splenectomized to exacerbate the pathological process. At the peak of parasitemia, the animal is totally bled. The resulting blood is canned and used for the manufacture of antigens. As a blood preservative, glugicir is used [1].

There is also a method according to which an animal is splenectomized to obtain blood for the production of antigen, then it is infected and at the peak of parasitemia, with damage to up to 50% of red blood cells, they completely bleed. The resulting blood is preserved with sodium citrate and used to produce antigen. This method is a prototype of the proposed [2].

The research objectives were to obtain a larger volume of full blood suitable for receiving highly active babesian antigens from one animal.

The proposed method is as follows. In a dog weighing 15-20 kg, the blood volume is approximately 2-2.5 liters. The animal is splenectomized. A few days after the operation, 150 ml of venous blood is taken from the animal, which is preserved with a glugicir in a ratio of 1: 4. Loss of blood is compensated by the introduction of isotonic solutions.

After 3-4 days, 300 ml of blood is taken from the animal, instead of which 150 ml of previously obtained blood and 150 ml of isotonic solution are administered. Then, after 3-4 days, 500 ml of blood is taken, which replace 300 ml of autologous blood and 200 ml of isotonic solution. Then after 3-4 days, 700 ml of blood is taken, which replace 500 ml of autologous blood and 200 ml of isotonic solution. And after 3-4 days they take 1000 ml of blood, which replace 700 ml of autologous blood and 300 ml of isotonic solution. The resulting blood is divided into two equal parts.

Then the animal is infected with the pathogen Babesia canis, the count of parasitemia is carried out by microscopy of blood smears stained according to Romanovsky-Giemsa. When parasitemia is more than 50%, 800-900 ml of blood is taken from the animal, which is filled up with 500 ml of prepared early blood (or half of the prepared blood) and 300-400 ml of isotonic solution. A decrease in parasitemia is noted. With its repeated growth, the procedure is repeated similarly to the first blood sampling. At the third peak of parasitemia, the animal is bled. Obtained at all three stages, the blood is canned with glucicir and used to produce antigen.

Duration of collection and blood volume may vary depending on the size of the animal. Specific examples are shown in the table.

Feasibility studies. The proposed method allows to obtain a larger volume of blood from one animal, despite longer lead times compared to the previous one. Infusion of non-invasive blood and isotonic solutions has a positive effect on the body of a sick dog.

The proposed method can be used to obtain blood for the preparation of antigens and from other types of babesias affecting other animals.

The proposed method was tested with a positive result in the veterinary clinics "Animal Park" and "Zoosphere" in 2007-2008.

This method will find application in the manufacture of babesian antigens in research institutes and veterinary production laboratories and will strengthen measures to combat blood parasitic diseases.

Information sources

1. Abstract of diss. ... candidate of veterinary sciences: Semenko OV The Meroia lively diagnosis of dog babies diagnosis, Kyiv, 2007 - 21 p.

2. Guidelines for the study and development of measures to combat animal protozoal diseases, Moscow, VASKHNIL, 1984 - p.7. (Prototype)

3. Abstract of diss. ... candidate of veterinary science: Kosheleva MI Babesiosis of dogs in the conditions of the Moscow region, Moscow, 2006 - 3 p.

Table The volume of blood obtained from animals with different weights, in comparison with the prototype The mass of the animal, kg Prototype, ml The proposed method, ml 10 500-600 1000-1400 twenty 1000-1500 2500-3000 thirty 1500-2000 3000-3500

Claims (1)

A method of obtaining blood for the manufacture of antigens from hemisporidia of Babesia canis, including splenectomy of an animal, infection with blood parasites, bleeding at the peak of invasion, preservation of blood and its use for the manufacture of antigens, characterized in that before infection, autologous blood is accumulated by five times taking and transfusion of autologous blood in the amount of 150 , 300, 500, 700 and 1000 ml, respectively, together with an isotonic solution with an interval of 3-4 days, after which the animal is infected with Babesia canis and at the peak of parasitemia with a lesion of 30-50% ritrotsitov take twice the blood for 800-900 ml, followed by infusion of autologous blood previously obtained, and then at the third peak of parasitaemia bled animal totally with 3-4 days interval, and the resulting three stages of the blood is used for the manufacture of antigen.