RU2558114C1 - Method for selecting donors of blood plasma applicable for treating patients with infectious diseases and postoperative, injury and burn complications - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: invention refers to medicine, namely to transfusion medicine, and is applicable in selecting blood plasma donors. That is ensured by sampling blood plasma to be analysed for antimicrobial peptides defensins. If the male defensin titre is from 180 to 545 ng/ml, and the female one is from 128 to 386 ng/ml, a donor is considered to be suitable for blood plasma drawing.

EFFECT: invention enables using the donor plasma in treating the patients suffering from infectious diseases and postoperative, injury and burn complications.

Description

The invention relates to medicine, in particular to blood transfusion, and can be used in the selection of donors to obtain blood plasma, effective in treating patients with infectious diseases and complications after surgery, injuries and burns.

There is a method of selecting donors by determining bacterial antibodies in their blood plasma, widely used by laboratory departments of blood transfusion stations (departments), in accordance with the acquisition plan and proposed treatment measures in institutions for the following infections with a significant therapeutic antibody titer:

- antistaphylococcal in a titer of 1: 1280 and above;

- anticolibacillary in a titer of 1: 160 and above;

- antiseptic in titer 1: 160 and higher;

- antiprotein in a titer of 1: 160 and above;

- anticlebsiellosis in a titer of 1: 160 and higher (A.V. Chechetkin et al., 2008 / Methodological guidelines “Obtaining and clinical use of immune plasma in military medical institutions”).

In the practice of industrial transfusiology, two solutions to the issue of providing immune plasma were used. This is a targeted immunization of donors and screening of antibacterial antibodies in donors, allowing you to select samples of freshly frozen plasma with antibodies with therapeutic titer (titers indicated above).

The main indicator of donor plasma sampling is the magnitude or significant therapeutic titer of antibacterial antibodies (AT) for transfusiological care. The percentage of AT detection in personnel donors ranged from 12 to 73% and depended on gender, age, season, and seasonal work. Control of plasma activity of donors with standard immune serum was carried out once every 3-4 weeks. Plasma samples are subsequently stored. If freezing is carried out within 6 hours after the blood supply, then this plasma is considered frozen (A.V. Chechetkin et al., 2007 / Method. Instructions - “Organization of quarantine freshly frozen plasma in military medical institutions”), and may also be lyophilized on time and stored in a dry condition. Its use is important in the field, as well as in remote areas from large medical institutions.

Since 2010, due to the lack of testing for the infections previously indicated by us, it prompted us to look for a new approach in the examination of donors in order to determine the natural antibacterial antibodies in plasma, which are essential for the provision of medical care and targeted use of donor plasma of a certain category of patients.

The search for highly effective means of therapy and prophylaxis of various infections for the blood service is currently gaining special urgency and urgent task of practical public health.

The use of a polyimmune plasma is advisable in case of a verified intrahospital infection (Pseudomonas aeruginosa, Staphylococcus aureus, Streptococcus, etc., pyogenic flora), a burn disease complicated by a secondary purulent infection, and in the clinic of internal diseases (severe bacterial pneumonia, septicemia, etc.).

The disadvantages of the known methods are the duration and difficulty of identifying donors, as well as the lack of a legal basis for the practical implementation in the clinic of the above treatment methods.

The aim of the invention is to increase the effectiveness of the treatment of patients with infectious diseases and complications after surgery, injuries and burns through the use of donor blood plasma containing antimicrobial peptides (defensins) that have microbicidal, chemotactic, immunomodulating and cytotoxic activity without intentional immunization of donors.

The goal is achieved by studying the immunological parameters of blood, and the content of antimicrobial peptides of defensins in the plasma is revealed and blood sampling is carried out in individuals whose plasma they contain at a titer of from 180 to 545 ng / ml in men, from 128 to 386 ng / ml in women.

The method is implemented as follows.

Investigations of immunological parameters of blood of donors are carried out, the content of antimicrobial peptides of defensins is determined and blood sampling for use in the treatment of seriously ill patients is carried out in individuals whose plasma they contain at a titer of 180 to 545 ng / ml in men, from 128 to 386 ng / ml - among women.

Practical example 1. Donor - I. (male) antimicrobial peptide titer was 550.6 ng / ml; blood plasma sent to the clinic of field surgery for the treatment of seriously wounded, delivered from Dagestan

Practical example 2. Donor - K. (male) antimicrobial peptide titer was 33.1 ng / ml; blood plasma is sent to the clinic of faculty therapy.

Practical example 3. Donor - L. (female), the titer of antimicrobial peptides amounted to - 228.9 ng / ml; blood plasma is sent to the clinic of field therapy for the treatment of a patient with severe poisoning.

Practical example 4. Donor - I. (female) antimicrobial peptide titer was 116.1 ng / ml; blood plasma is sent to the clinic of obstetrics and gynecology.

Currently, antimicrobial peptides are an object of increased attention of a wide range of specialists, including those considering such compounds as an alternative to antibiotic agents. To counter pathogens, scientists are developing more and more types of antibiotics, which are essentially derivatives of the old ones. Antimicrobial peptides are inferior to antibiotics in effectiveness, but act faster, destroying antibiotic-resistant bacteria (Garsia A, E, Osapau G, Tran PA, Yuan J, Selsted ME Infect Immun. 2008 Dec. 76 (12): 5883-91).

Peptides are constantly formed even in a stable state of the body. A.E. Abaturov. 2012 Child Health, 3 (38) 2012, indicates that even at low concentrations of defensins inhibit adenovirus infection.

Antimicrobial peptides-defensins are multifunctional cationic peptides of humans and are short molecules with a length of 12 to 50 amino acids, have an average molecular weight of 4 kD, perform complex functions in maintaining and regulating the immune response to viral, microbial and other antigenic aggression based on direct effects on the cell membrane of the microorganism that causes cell destruction, as well as participation in the regulation of antigen presentation.

Antimicrobial antibody-based drugs have not yet been created. At the same time, indicators of the content of defensins in blood plasma (human neutrophil protein) is considered as an indicator of the presence of high immunoprotective function.

B.V. Pinegin, A.S. Budikhina in the journal "Immunology" 2008, No. 5, pp. 317-320 indicate that antimicrobial peptides (defensins) are components of the immune system and provide the first line of defense of a macroorganism from various pathogens.

1. Advantages of the method:

- study time 3.5 hours;

- the minimum measured concentration of 156 pg / ml;

- the measured range of 156-10000 pg / ml;

- sample volume 100 μl per well;

- the availability of test systems for research.

2. The principle of the method.

It is based on the "sandwich" method of enzyme-linked immunosorbent assay using the diagnostic kit HNP 1-3 (neutrophil defensins) ELISA.

2. The preparation of blood samples of donors for research requires the strictest implementation of all the recommendations of the methodological instructions.

We examined 88 personnel donors aged 21-34 years, of which 81 were males, 7 were women.

The obtained blood samples of donors were stored at + 2-4 ° C. Plasma was separated by centrifugation no later than 20 minutes after blood exfusion at 1500 g at + 4 ° C for 15 minutes. Plasma was collected in clean polypropylene tubes, carefully, without touching the cell pellet. Re-centrifuged under the same conditions. The most reliable results were obtained using heparin or EDTA as an anticoagulant.

Samples were stored at -70 ° C in polypropylene tubes prior to testing. Storage at -20 ° C negatively affects the recovery of HNP 1-3 defensins.

Studies of donor blood samples were carried out 24 hours after thawing and preliminary dilution with the supplied buffer and mixing in polypropylene tubes.

The preparation of reagents, the study procedure was carried out in accordance with the instructions.

Of the 81 men examined, 20 revealed HNP 1-3 defensins from 180 to 545 ng / ml - 24.7%, in 2 women - from 128 to 386 ng / ml - 28.5%.

Claims (1)

A method of selecting donors for obtaining blood plasma by studying immunological blood parameters, characterized in that the plasma reveals the content of antimicrobial peptides of defensins and with a defensin titer of 180 to 545 ng / ml in men and from 128 to 386 ng / ml in women blood sampling.