Method of determining individual human blood leukocyte drug sensitivity

FIELD: medicine.

SUBSTANCE: method of determining individual human drug sensitivity or immunotoxicity is based on individual modelling in vitro of the effect on IFN-alpha and/or IFN-gamma production by patient's whole blood leukocytes/lymphocytes. The method involves evaluating a degree of a stimulating/correcting or inhibitory drug action and/or afteraction on the primary IFN system reactivity values.

EFFECT: method provides individual prescription of adequate drugs and thereby higher clinical efficacy for prevention of potential developing medicinal interferon immunodeficiencies.

3 cl, 4 ex

 

The invention relates to medicine, namely to immunology, pharmacology, Virology, and the development of a method for determining individual sensitivity of leukocytes/lymphocytes human blood to immunotropic or other drugs, modifying, depending on such, in varying degrees, the production of immunocompetent cells interferon (IFN) - the main anti-infective cytokines, the level of production which reflects the intensity antiviral and antibacterial immune responses of the organism as a whole.

The method can be used to predict clinical efficacy (or possible immunotoxicity) immunotropic (or other drugs) drugs and their scientifically individual selection for therapy of various diseases.

Currently, the pharmaceutical market offers a wide Arsenal of immunotropic and/or immunoactive drugs: recombinant and natural human interferon, interleukins, inducers of IFN and immunomodulators used in the prevention and treatment of viral and other infectious, allergic, autoimmune, cancer and various immunopositive diseases. The response to these drugs in the human population, depending on the individual characteristics of each organism, the nature of the RA and severity of the pathological process, may vary significantly and can be expressed not only in the presence of the expected positive immune correcting action and clinical effect, but in the absence of such, or, more importantly, to reverse the suppressive effects and unwanted side effects.

In addition, a large Arsenal of various pharmacological agents used in medical practice, is not considered formally to modulate immune system activity or synthesis of cytokines (corticosteroids, antibiotics, sulfonamides, anti-TB and other drugs), can, along with the main pharmacological action adversely affect IFN-response of blood leukocytes to cause suppression of the IFN system, cellular and humoral immunity and, therefore, to provide overall immunotoxic effect.

The technical purpose of this invention to provide advance information about possible immunokorrigiruyuschy the effect of the drug on blood leukocytes of the patient and, therefore, about its individual sensitivity to immunotropic drugs and portability of their other medicines and related forecasting their subsequent clinical effectiveness, allowing science to justify and facilitate the adequate choice for this patient immunoactive drug, sboev the temporal prevent and correct possible immunotoxicity of other medicines.

To solve this problem is individual modeling of in vitro exposure to various immunotropic or immunoactive (or other drug) drugs in whole blood of the patient, which is estimated by the change under action 2 of the main indicators of IFN-status: IFN-α and IFN-γ producing capacity of the blood leukocytes of the patient, i.e. to change the number produced by leukocytes/lymphocytes his blood IFN-α and IFN-γ in vitro, costimulatory each of the study drug.

The prototype of the proposed method is developed by us earlier method of determining IFN-status people in samples of whole blood (1-3)including testing of the following indicators:

- the amount of circulating blood IFN (serum IFN),

- level of production by leukocytes/lymphocytes whole blood IFN-α-induced in vitro reference inducer of IFN-α - virus Newcastle disease (VBN),

- level of production by leukocytes/lymphocytes whole blood IFN-γ induced in vitro reference inducer of IFN-γ phytohemagglutinin (PHA),

- level of production by leukocytes/lymphocytes whole blood spontaneous IFN in vitro without any additional induction.

The present invention differs from the prototype that is tested and compared quantitative content produc is one of the leukocytes/lymphocytes whole blood of the patient in vitro IFN-α and IFN-γ under the action of one or the other immunoactive or other pharmaceutical without it.

In other words, simulated and evaluated individual co-stimulating/corrective effects of drugs on 2 of the most informative indicator of interferonogenesis and reactivity of the IFN system of the patient-mediated sensitivity of leukocytes/lymphocytes his blood to each of the study drug, i.e. is determined by the individual index costimulation and correction of each drug for a given patient.

Depending on the index costimulation and correction (index QC) products of Itnow α and γ by leukocytes/lymphocytes whole blood in vitro possible 5 main options individual sensitivity (ICH) to immunoactive (inducers of interferon, immunomodulators, cytokines) and other drugs:

1. Weak ICH - (index QC=2) is 2-fold increase in titers of IFN-α and IFN-γ induced in vitro in the presence of the drug compared with those in the absence of the drug in the standard induction (figures production of IFN-α and IFN-γ according to IFN-status of the patient).

2. Expressed ICH - (index QC=3-4) - 3-4-fold increase in titers of IFN-α and IFN-γ induced in vitro in the presence of the drug compared with those in the absence of the drug in the standard induction (figures production of IFN-α and IFN-γ according to IFN-status of the patient).

3. Strongly expressed ICH - (index QC>4) - Bo is it than the 4-fold increase in titers of IFN-α and IFN-γ, induced in vitro in the presence of the drug compared with those in the absence of the drug in the standard induction (figures production of IFN-α and IFN-γ according to IFN-status of the patient).

4. The absence of ICH - index (QC=1) - titers of IFN-α and IFN-γ induced in vitro in the presence of the drug compared with those in the absence of the drug in the standard induction (figures production of IFN-α and IFN-γ according to IFN-status of the patient), remain unchanged.

5. Individual immunotoxicity (AI) - (index QC<1) titers of IFN-α and IFN-γ induced in vitro in the presence of the drug lower than those in the absence of the drug in the standard induction (figures production of IFN-α and IFN-γ according to IFN-status of the patient).

It is known that the system of IFN man is the first line of resistance of the organism to infection and its immunoreactivity in other forms of pathology.

According to modern concepts, immunotropic or immunoactive drugs exert their effects on the system of IFN person through the induction of not only (and sometimes not so much) IFN, but also a number of other Pro - and anti-inflammatory cytokines, providing, effective forward and backward linkages, positive or negative impact on interferonogenez in General, and α - and γ-IFN producing ability of leukocytes/lymphocytes in particular.

Other f mikologicheskie drugs not related to immunotropic or immunoactive can also inhibit IFN-response of immunocompetent blood cells (including as a result of changes or damage signaling pathways of these cells and, hence, to suppress the resistance and the immunoreactivity of the organism as a whole.

The inventive method allows us to evaluate and predict individual tolerability and potential immunotoxicity of these drugs, to prevent its adequate for this patient immunotropic drug.

Staging tests on ICH and tolerability of drugs by the present method may be carried out in 3 versions:

Option I - the effect of the investigational product, when all reactive components: the blood of the patient, the drug at a final concentration corresponding to its one-time and/or daily therapeutic dose, and the reference inducer of IFN-α or IFN-γ are introduced into the reaction mixture simultaneously, i.e. the reaction is carried out in the presence of the drug.

Option II - the aftereffect of the investigational product, when is the 24 hour pre-incubation of the blood of a patient with an investigational drug in a final concentration corresponding to its one-time and/or a daily dose, with subsequent introduction of standard inductors α - and γ-Itnow.

Option III - the effect of the tested preparation is the action and/or delay) is measured by the results of its impact only on the γ-point of the IFN system, i.e. the production by leukocytes/lymphocytes of a patient's blood in vitro only IFN-γ is a key cytokine that defines the direction and intensity of the immune response during secondary immunodeficiencies associated with viral and other infectious, autoimmune, cancer and various immunopositivity diseases.

All 3 versions along with production tests ICH conducted a study of IFN-status 4-m major indices (1-3), which allows to evaluate the functional activity of the IFN system in situ in General and to determine the degree of failure on the various links in this patient with this disease.

The failure of the IFN system in α - or γ-links (IFN-α or IFN-γ-producing ability of leukocytes/lymphocytes) can be:

1st degree - when indicators production of IFN-α or IFN-γ by leukocytes/lymphocytes according to the determination of IFN-status lower than those in the norm 2 times,

2-nd degree - when indicators production of IFN-α or IFN-γ by leukocytes/lymphocytes according to the determination of IFN-status lower than those in the normal 4 times,

3rd degree - when indicators production of IFN-α or IFN-γ by leukocytes/lymphocytes according to the determination of IFN-status lower than those in the normal 8 times,

4th degree - when indicators production of IFN-α or IFN-γ by leukocytes/lymph what it blood according to the determination of IFN-status lower than those in the norm more than 8 times.

Comparison of indicators of production of Itnow-α and/or γ options I, II or III with the control (according to IFN-status, without preparation) allows to judge the presence or absence and the degree of co-stimulating/corrective (or inhibition) of action of this drug on the corresponding indicators of the patient (indexes QC or AI of the drug), and hence of ICH or endurance by leukocytes/lymphocytes his blood investigational product.

Activity-induced α - and/or γ-Itnow in all variants is determined after 24 hours of contact with a reference inducers of IFN-α and IFN-γ biological or enzyme immunoassay method (ELISA) using diploid cultures of human fibroblasts or appropriate test systems and is expressed in IU/ml or PCG/ml (picograms/ml), respectively.

Options I, II and III as appropriate in the study of immunotropic/immunoactive and other drugs. In practice, often, usually used in test performance on ICH for Option III.

No ICH patient to a particular drug does not currently preclude its recovery in subsequent during treatment or after cure of this disease, as well as available at the beginning of the disease high sensitivity to this drug in the course of treatment may be reduced. Therefore, when licenciamento/immunoactive drugs, it is advisable to monitor ICH patient, that will allow time to cancel one and apply another drug, i.e., to exercise reasonable combination therapy of various immunotropic/immunoactive drugs to achieve maximum clinical effect. You can also increase the concentration of drugs within the maximum allowable disposable therapeutic doses. In such cases, the response to ICH are carried out in the presence of various calculated concentrations of the same drug.

Examples of procedures for carrying out the invention.

Example 1. The definition of ICH leukocytes/lymphocytes of the patient K., 28 years old, sickly (5-6 times a year) ARVI, inducers of IFN - Amiksin and cold and immunomodulators - polyoxidonium and Likopid (the choice of the treating physician) - only 4 of the drug.

Blood taken from a vein (or ring finger) any hand after pre-processing first alcohol and then a dry cotton swab, sterile disposable syringe (or scarificator) in a volume of 1-2 ml and transferred into a sterile plastic microprobing containing 10-20 units of heparin or other anticoagulant.

Option I - the effect of the study drug.

Staging tests ICH drugs carried out in parallel with the definition of the indicators of IFN-status of the patient without the drug 3 times in total volume of 200 m is l all reactive components, including the patient's blood.

In sterile 96-well round-bottom plate make the medium RPMI-1640 (160 μl in 9 holes and 140 cells / ml in 24 wells.

In each well, containing 160 μl of RPMI, make:

1) in the first three holes 20 µl VBN (reference inducer of IFN-α),

2) in the second three holes 20 μl of PHA (reference inducer of IFN-γ),

3) in the third three holes 20 µl of RPMI-1640 medium (spontaneous induction of IFN).

To each well containing 140 μl of RPMI-1640, make:

4) in the first 6 holes 20 µl of the drug Amiksin in a single therapeutic dose + 20 µl VBN 3 wells (costimulate production of IFN-α in the presence of Amiksin) and 20 μl of PHA in 3 other wells (costimulate production of IFN-γ in the presence of Amiksin);

5) in the second 6 holes 20 µl of the drug cycloferon in a single therapeutic dose + 20 µl VBN 3 wells from them and 20 μl of PHA in 3 other wells (costimulate products of Itnow α and γ, respectively, in the presence of cycloferon);

6) in the third 6 holes 20 µl of the drug polyoxidonium in a single therapeutic dose + 20 µl VBN 3 wells from them and 20 μl of PHA in 3 other wells (costimulate products of Itnow α and γ, respectively, in the presence of polyoxidonium);

7) in the fourth 6 holes 20 µl of the drug Likopid in a single therapeutic dose + 20 µl VBN 3 of them and 20 μl of PHA in 3 other (costimulate products Ifno α and γ, respectively, in the presence of licopid);

8) in each of the holes with reaction mixture (for the version I just 33 wells) add 20 μl of heparinised whole blood analyzed patient;

9) the contents of each well, mixed well, the tablets covered with a lid and incubated at 37°C in an atmosphere of CO224 hours;

10) after 24 hours incubation tablets centrifuged in normal mode, the supernatant liquid in a volume of 150 ál from each well of the 3 repetitions are selected and combined into one sample (a total of 11 samples).

The flow of blood in the described procedure productions ICH and IFN-status is 660 mm. The remaining blood is centrifuged to obtain and determine the content of serum IFN patient.

Titration of biological activity of the obtained samples IFN spend in diploid culture of human fibroblasts in 3 repetitions or determine the content in the studied samples of IFN-α and IFN-γ in 3 repetitions using the appropriate test systems conventional methods, which are expressed in IU/ml or PCG/ml, respectively.

Example 2. Option II - the aftereffect of the studied drugs.

The procedure of registration ICH leukocytes/lymphocytes of the patient K. drugs Amiksin, cycloferon, polyoxidonium and Likopid differs from the previous version that previously, within 24 hours, contact the studied drugs with kr the view of the patient (without reference inducers of IFN-α and IFN-γ).

In sterile 96-well round-bottom plate is made the medium RPMI-1640 to 180 ál in 9 holes and 160 cells / ml in 24 wells;

In each well, containing 160 μl of RPMI-1640, make:

1) paragraphs 4, 5, 6, and 7 option I - only 20 µl of drugs Amiksin, cycloferon, polyoxidonium and Likopid, respectively, 6 holes on each drug in pay and/or daily therapeutic dose (24 wells);

2) in each of the holes with reaction mixture (i.e. in all 33 wells on items 1-7 option I) add 20 μl of heparinised whole blood analyzed patient;

3) the contents of each well, mixed well, the tablets covered with a lid and incubated at 37°C in an atmosphere of CO224 hours;

4) after 24 hours incubation tablets centrifuged in normal mode, the supernatant liquid is poured in the wells contribute 20 µl of VBN and PHA in 3 repetitions, paragraphs 1, 2, 4-7 options I and 20 µl of RPMI-1640 medium under item 3.

5) the contents of each well, mixed well, the tablets covered with a lid and re-incubated at 37°C in an atmosphere of CO224 hours;

6) then follow the procedure in paragraph 10 of option I and determine the activity induced by IFN similarly to the previous example.

Options I and II productions of ICH to investigational drugs can be delivered separately or in parallel.

Example 3. Option III - effect of action and/or aftereffect and the subjugated drugs on the production by leukocytes/lymphocytes of a patient's blood IFN-γ.

The procedure of registration ICH leukocytes/lymphocytes of the patient K. drugs Amiksin, cycloferon, polyoxidonium and Likopid under this scenario include:

Sub-option A - the effect of the study drug.

In sterile 96-well round-bottom plate is made the medium RPMI-1640 160 cells / ml in 9 holes and 140 cells / ml in 12 holes;

To each well containing 160 μl of RPMI-1640, make:

1) 20 µl VBN, PHA or RPMI-1640 medium in 3 repetitions, respectively (3 wells each) according to paragraphs 1 to 3 of option I (for staging tests IFN-status);

To each well containing 140 μl of RPMI-1640 medium, make:

2) 20 μl of each of the investigated drugs in 3 times (3 wells for each drug in a single therapeutic dose) + 20 ál PHA in all 12 holes;

3) all wells with reaction mixture (9+12 - a total of 21 wells) is added 20 μl of heparinised whole blood analyzed patient;

4) the contents of each well, mixed well, the tablets covered with a lid and incubated at 37°C in an atmosphere of CO224 hours;

5) after 24 hours incubation tablets centrifuged in normal mode, the supernatant liquid in a volume of 150 ál from each of the 3 repetitions combined into one sample (7 samples);

6) activity induced by Itnow in the obtained samples samples determined as described in the previous versions.

Sub-option is - the aftereffect of the studied drugs.

In sterile 96-well round-bottom plate is made the medium RPMI-1640 to 180 ál - in 9 holes and 160 cells / ml in 12 holes;

In each well, containing 160 μl of RPMI-1640 medium, make:

1) 20 µl of each of the studied drugs in 3 times (3 wells for each drug in pay and/or daily therapeutic dose);

2) all wells with reaction mixture (9+12 - a total of 21 wells) add 20 μl of heparinised whole blood analyzed patient;

3) the contents of each well, mixed well, the tablets covered with a lid and incubated at 37°C in an atmosphere of CO224 hours;

4) after 24 hours incubation tablets centrifuged in normal mode, the supernatant liquid from all wells is poured in the wells contribute 20 µl VBN, PHA or RPMI-1640 medium in 3 repetitions, respectively (3 holes on items 1-3 options 1-9 holes for staging tests IFN-status), all the other 12 holes, the preprocessed investigational drugs, contribute 20 ál PHA;

5) the contents of each well, mixed well, the tablets covered with a lid and re-incubated at 37°C in an atmosphere of CO224 hours;

6) after re-incubation tablets centrifuged in normal mode, the supernatant liquid in a volume of 150 ál from each of the 3 repetitions combined into one sample (7 samples);

7) Akti the ability of induced Itnow in the obtained samples samples define as described in the previous versions.

The sub-options a and B Option III staging tests ICH to investigational drugs can be delivered separately or in parallel.

Examples of results and conclusions on the definition of ICH to different drugs.

Example 1 - patient Option K. I.

Conclusion.

According to a study in example 1 in a patient K., deficiency of IFN system by production of IFN-alpha and IFN-gamma 2-nd and 3-rd degree, respectively. Is determined by a pronounced sensitivity to cold in costimulation and correction of production of IFN-gamma (index QC is equal to 4) and weakly expressed in costimulation and correction of production of IFN-alpha (index QC equal to 2). To the other tested drugs (polyoxidonium, Likopid) leukocytes/lymphocytes of the patient K. less sensitive (indexes QC production of IFN-alpha or IFN-gamma, respectively equal to 2) or not sensitive (Amiksin).

It is recommended that treatment with cycloferon, whose action on the alpha-link of the IFN system may be enhanced by combination with polyoxidonium.

Example 2 - the patient is N. Option II.

Conclusion.

According to a study in example 2, the patient H. there is a failure of the IFN system by production of IFN-alpha and IFN-gamma 3rd degree, respectively. Determined the tsya strongly expressed sensitivity to Radostina on costimulation and correction of production of IFN-alpha (index QC is equal to 8) and expressed in costimulation and correction of production of IFN-gamma (index QC is equal to 4), and severe sensitivity to cold in costimulation and correction products as IFN-alpha and IFN-gamma (index QC is equal to 4). To the other tested drugs (Imunofan, polyoxidonium) leukocytes/lymphocytes of the patient H. after 24 h incubation, less sensitive (indexes QC on the production of IFN-alpha or IFN-gamma, respectively equal to 2) or not sensitive (polyoxidonium).

Recommended treatment Radostina, cycloferon, the possible combination of cold + Imunofan.

Example 3 - the patient C. Option III (sub-options a and B).

Conclusion.

According to a study in example 3, the patient With. there is a failure of the IFN system by production of IFN-alpha and IFN-gamma 2-nd and 4-th degree, respectively. Is determined by the strongly expressed the sensitivity of leukocytes/lymphocytes of a patient's blood to Radostina on costimulation and correction of production of IFN-gamma as under the influence, and after his actions (index QC is equal to 8 and 16 respectively), as well as a pronounced sensitivity to cold in the presence of and after which the production of IFN-gamma increased at 4 and 8 times, respectively, and Imunofan, effect and aftereffect of which will costimulated IFN-gamma-producing ability immunocompetent who's blood cells of the patient 4 times. The less pronounced sensitivity is determined by the action of neovir, which, however, after 24 hours of contact with the blood of the patient also is expressed costimulation production of IFN-gamma by leukocytes/lymphocytes of a patient's blood S. (index costimulation equal to 4).

Recommended treatment Radostina, cycloferon, Imunofan, or a combination of neovir + Imunofan.

Example 4. The patient So the definition of the potential immunotoxicity of antibiotics, cephalexin, doxycycline and cytostatic cyclophosphamide and 5-fluorouracil. Option III (sub-options a and B).

Conclusion.

According to a study from the patient So there is a lack (deficiency) of IFN system by production of IFN-alpha and IFN-gamma 4-th degree. Determined moderate immunotoxic effects of cephalexin on the production of IFN-gamma by leukocytes/lymphocytes of a patient's blood (A), which increases after 24-hour incubation with the drug (B) and expressed immunotoxic effects of 5-fluorouracil on the production of IFN-gamma (B).

In the treatment of various diseases for the patient So it is preferable application of the antibiotic doxycycline and cytostatic cyclophosphamide.

In General, the determination of individual sensitivity and immunotoxicity of various drugs by the present method aims at getting ahead in the information about the clinical effectiveness of immunotropic/immunoactive drugs and preventing the possible development of drug interferon immune deficiencies and can be used as an important additional test preclinical studies of a wide range of pharmacological preparations.

Literature.

1. Grigoryan, S., majors, I.A., Ivanov, A.M., Ershov FI Evaluation of interferon status of people on samples of whole blood. Questions Virology 1988, 4, 433-436.

2. Grigoryan, S., Ivanov, A.M., Pritzker A.D., Ershov FI Determination of interferon status in whole blood from people for mass screening. M., Academy of medical Sciences of the USSR, 1989, p.1-14.

3. Grigoryan, S., Ershov FI Methodological principles for determining the interferon status. In kN. "The interferon system in normal and pathological conditions". M, Medicine, 1996, str-155.

1. The method of determining individual sensitivity (ICH) of blood leukocytes to medicinal drug, characterized in that after 24 hours and/or after the action of the drug in whole blood of the patient in vitro in pay and/or daily dose to increase or reduce the level of production (subtitles) IFN-alpha and/or IFN-gamma by peripheral blood leukocytes, compared with those in the absence of the drug, determine the degree of co-stimulating or suppressing the action of the drug on these indicators of the patient; calculate the index costimulation and correction (QC) of the drug and assess ICH to medicinal drug, dividing by 5 main options: weak ICH - 2-fold increase titro the IFN-alpha and/or IFN-gamma and index QC=2; expressed ICH - by 3-4-fold increase in titers of IFN-alpha and/or IFN-gamma and index QC=3-4; highly expressed ICH - when more than 4-fold increase in titers of IFN-alpha and/or IFN-gamma and index QC>4; lack of ICH - in the absence of changes in the titers of IFN-alpha and/or IFN-gamma and index KK=1; private immunotoxicity (AI) - at lower titers of IFN-alpha and/or IFN-gamma and index QC<1.

2. The method according to claim 1, in which the induction of IFN-alpha and/or IFN-gamma in leukocytes whole blood of the patient corresponding reference inducers in vitro is carried out in the presence of the drug in pay and/or daily dose (effect of drug action).

3. The method according to claim 1, in which the induction of IFN-alpha and/or IFN-gamma in leukocytes whole blood of the patient corresponding reference inducers in vitro is carried out 24 h after drug action in pay and/or daily dose (the aftereffect of the drug).



 

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2 ex

FIELD: medicine, medicinal microbiology.

SUBSTANCE: method involves growing microorganism culture to be studied in solid nutrient medium followed by preparing microbial suspension and its incubation in the presence of lactoferrin. Control sample is prepared in parallel series. Control and experimental samples are incubated, supernatant is removed from bacterial cells and lactoferrin concentration is determined in supernatant of experimental and control sample by immunoenzyme analysis. Then anti-lactoferrin activity is calculated by difference of concentrations of residual lactoferrin in experimental and control samples. This method provides enhancing the sensitivity and precision in carrying out the quantitative evaluation of anti-lactoferrin activity in broad spectrum of microorganisms that is urgent in diagnosis and prognosis of diseases with bacterial etiology. Invention can be used in determination of persistent indices of microorganisms for assay of their etiological significance in pathological processes.

EFFECT: improved assay method.

3 tbl, 3 ex

FIELD: medicine, biology.

SUBSTANCE: invention relates to nutrient medium used for accumulation of cells for the following cytological and/or immunocytochemical analysis carrying out. Invention relates to medium containing salts NaCl, KCl, anhydrous CaCl2, MgSO4 x 6 H2O, MgCl2 x 6 H2O, Na2HPO4 x 2 H2O, KHPO4, NaHCO3, and also glucose and Henx's solution, 10% albumin solution and polyglucin taken in the ratio 1:1:1. Invention provides enhancing the preservation of cells.

EFFECT: improved an valuable properties of nutrient medium.

3 ex

FIELD: medicine, cardiology.

SUBSTANCE: in peripheral blood one should detect the level of CD95(+) and CD16(+) neutrophilic granulocytes and at combination of increased level of CD95(+) neutrophilic granulocytes by 4 times and more and CD16(+) neutrophilic granulocytes by 0.6 times against the norm with ECG signs of myocardial infarction one should predict lethal result of large-focal myocardial infarction.

EFFECT: higher accuracy of prediction.

FIELD: medicine, parasitology.

SUBSTANCE: one should carry out immunoenzymatic assay to detect diagnostic optic density and that of labeled immune complex in a plot's hole with tested serum measured in conventional units at wave length being 492 nm. One should calculate coefficient of antibodies concentration measured in conventional units by the following formula: CAC = (Odtsh - Odd) x 100, where CAC - coefficient of antibodies concentration, Odtsh - optic density of the hole with tested serum, Odd - diagnostic value of optic density, 100 - coefficient of serumal dilution. By CAC value one should detect the titer of antibodies to Lamblia intestinalis antigens to interpret results of the trial. The method enables to study the dynamics of disease flow.

EFFECT: higher efficiency and accuracy of diagnostics.

1 ex, 1 tbl

FIELD: medicine.

SUBSTANCE: the present innovation deals with studying and treating diseases of inflammatory, autoimmune and degenerative genesis. One should perform sampling of heparinized blood followed by its sedimentation to obtain blood plasma with leukocytes and centrifuging to isolate the latter which are washed against erythrocytic and serumal admixtures, and, also, it deals with calculating the number of cells in samples out of leukocytic suspension after incubation (B) for 1.5 h at 37 C in holes of plastic microplotting board, out of leukocytic suspension one should additionally prepare two samples, one should be applied to calculate total number of leukocytes before incubation (A), the second sample undergoes incubation at the same mode at addition of autoserum to calculate the number of cells remained after incubation (C). One should state upon adhesive properties of leukocytes by the index of spontaneous adhesion (D), where D=(A-B)/B.100%, and effect for enhanced cellular adhesion under the impact of autoserum should be detected by the value of K=(B-C)/C.100% at K ≥ 30%, where B - C - the number of cells undergone additional adhesion after addition of autoserum. The present innovation widens functional possibilities of the suggested method due to obtaining additional values depicting adhesive properties of blood leukocytes.

EFFECT: higher accuracy of detection.

FIELD: medicine, immunology.

SUBSTANCE: one should carry out reaction of blast-transformation, detect proliferation of T-lymphocytes activated with antibodies to CD3 in the presence of interleukin-7 (ACT IL-7) and in the presence of interleukin-7 and dexametazone (ACT IL-7 D), calculate the index for dexametazone action as the ratio of ACT IL-7 to ACT IL-7 D, moreover, the value of dexametazone action index being above 1.2 indicates increased production of cytokins that suppress T-lymphocytes in neonatals. The method enables to detect functional defect of immune system that characterizes neonatal period.

EFFECT: higher efficiency of detection.

2 ex

FIELD: medicine.

SUBSTANCE: method involves measuring forced exhalation volume per 1 s (FEV1) in l, full right ventricle evacuation time (RVE) in ms and angiotensin II value (AII) in ng/l. Discriminant relationship is built as D=0.504·RVE+3.038·FEV1 - 2.0·AII. D being less than 83.88, pulmonary hypertension occurrence is predicted within 1 year. D being equal to or greater than 83.88, no pulmonary hypertension is predicted to occur.

EFFECT: enhanced accuracy of prediction.

FIELD: medicine, medicinal immunology.

SUBSTANCE: method involves determination of heterophilic antibodies in human serum blood by the Paul-Bunnel's method relatively the level of circulating immune complexes, complement-activating properties of heterophilic antibodies by incubation of standardized ram erythrocytes with 0.8% serum for 30 ± 5 min and the following measurement of the erythrocytes lysis degree. The measurement of the effector function coefficient of heterophilic antibodies is carried out by the complement system Keff.f.h.a.-c.s. by the formula: Keff.f.h.a.-c.s. = Y/Tg.a. wherein Y means a lysis degree, %; Tg.a. means a reverse titer of heterophilic antibodies to ram erythrocytes. The damage assay is carried out by comparison of the immune status with the relative level of circulating immune complexes in serum. Method provides detection of preclinic from of immunodeficiency and autoimmune diseases that opens the possibility for their prophylaxis at most early stages of development. Invention can be used for assay of damage in the immune status in human serum blood.

EFFECT: improved method for assay.

5 tbl, 1 ex

FIELD: medicine.

SUBSTANCE: method involves concurrently examining anti-inflammatory IL-4 level in blood serum and lacrimal fluid. The value being within the limits of 60-70 pg/l in blood serum and 5-15 pg/l in lacrimal fluid, disease prognosis is considered to be unfavorable. The IL-4 concentration being within the limits of 90-100 pg/l in blood serum and 20-30 pg/l in lacrimal fluid, disease prognosis is considered to be favorable.

EFFECT: high accuracy of diagnosis.

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