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1.
The biological activities of tuberculin PPD RT 23 and the International Standard for Purified Protein Derivative of Mammalian Tuberculin (PPD-M) were compared in sensitized and unsensitized guinea-pigs by skin tests and lymphocyte stimulation (LS) tests. Estimates of relative potency (RP) from skin test results were dependent on the dose level, on the immunogen used, and, in guinea-pigs immunized with killed tubercle bacilli in oil, also on the immunization time. Relative potency estimates from LS results were dependent on the source of the lymphocytes and were different from estimates obtained from skin tests. Lymphocyte stimulation dose-response curves for the tuberculins were qualitatively different. In contrast to RT 23, PPD-M gave rise to non-specific skin reaction in unsensitized guinea-pigs. Both tuberculins were mitogenic to lymph node lymphocytes isolated from unsensitized guinea-pigs, PPD-M being the more mitogenic of the two tuberculins. The present results confirm that qualitatively different tuberculins cannot be unambiguously calibrated in identical terms and thus emphasize that the uncritical use of (international) standards should be avoided in tuberculin calibration.  相似文献   

2.
The modified leukocyte migration test (LMT) from an agarose droplet with the antigen stimulation (by BCG) is proposed in the present work. The BCG concentrations ranging from 1.25 up to 130 mg/l were used to examine 20 tuberculin (PPD) skin test negative and 10 PPD positive patients, which suffered from lung diseases. The optimal concentrations were 6.25 and 25.0 mg/l. The mean index values of a 20 membered control group ranged from 0.7 up to 0.88 when stimulated with the lower BCG concentration, and they were of 0.53 up to 0.69 at higher BCG concentration. In spite of its suitable indicatory properties as to ascertain cell mediated immunity state, the LMT with the BCG were of no use as a tuberculin skin test correlate.  相似文献   

3.
Wide heterogeneity was shown among different batches of bovine protein purified derivative (PPD) tuberculin, as regards protein content and antigenic profile. These features were also investigated in pilot preparations of Mycobacterium bovis secreted antigens and PPD tuberculins. Under controlled conditions, widely different compositions were revealed as a function of the M. bovis strain and also of the time in culture, due to the transient expression of seemingly important clusters of antigens. Furthermore, these parameters could dramatically affect the efficacy of the above preparations in in vitro assays of cell-mediated immunity on M. bovis-infected cattle. The field exposure to mycobacteria of the avium/intracellular group could also influence the readout of such assays, results being in agreement with a bystander suppression model of the response to M. bovis antigens. Due to the above, practical suggestions are put forward to improve the composition of bovine PPD tuberculins and the relevant control procedures.  相似文献   

4.
This study, conducted in 2009, proposed to evaluate and compare the biological potency of two different tuberculins, RT23 (Statens Serum Institute, Copenhagen) and IC-65 (Cantacuzino Institute, Bucharest) when administered to 89 children with confirmed tuberculosis, admitted to Paediatric Department of Pneumophtysiology Institute, Bucharest. Mean age of subjects was 10.4 years [SD (standard deviation) = 5.2 years; variance = 27.2], and sex distribution in the group was: 55.1% girls and 44.9% boys. Tuberculin skin tests were performed using Mantoux method simultaneously with the two tuberculins in the same concentration, 2TU (tuberculin units)/0.1 ml. RT23 skin test reactions ranged from 8 mm to 18 mm (mean = 12.8 mm, SD = 2.1 mm, variance = 4.4; median = 12.0), and IC-65 reactions ranged from 8 mm to 18 mm (mean = 13.1 mm; SD = 2.1 mm; variance = 4.3; median = 13.0). The mean difference in paired reaction sizes for the two reagents was 0.04 mm and was not statistically different from zero (P value = 0.3). The difference in reaction sizes was = 2 mm in 70.8% and = 5 mm in 7.9% patients. With a cutoff of 10 mm to define a positive reaction, the results were highly correlated with a sensitivity of 98.9% for RT23 and 97.8% for IC-65. No statistically significant difference was established for the efficacy of the two commercially available PPD TST reagents, both tuberculins appearing to have equivalent potency.  相似文献   

5.
Study on systemic reaction of delayed type hypersensitivity   总被引:1,自引:0,他引:1  
Effectiveness of two different samples of PPD tuberculins was studied quantitatively. No differences were found in the effect on skin test in rabbits, extent of skin reaction in guinea pigs, edema of foot-pad in rats and inhibition of spleen cell migration in guinea pigs. Differences were observed in the systemic febrile reaction in rabbits and in examinations of the thickness of infiltrated skin in guinea pig tests. The results may serve as a basis for standardization of systemic reaction in rabbits.  相似文献   

6.
A comparison of the direct migration inhibition using peritoneal exudate cells and peripheral blood leucocytes from the same tuberculin sensitive guinea pigs was performed. Three antigen concentrations: 3, 15, and 75 microgram of PPD per ml were used. Both type of cells provided similar results except at early incubation intervals when leucocytes, in the presence of lower doses of the antigen, displayed stronger inhibition than peritoneal cells. Thus, in our study, peripheral blood leucocytes are at least equivalent to peritoneal exudate cells in the migration inhibition test.  相似文献   

7.
Tuberculin purified protein derivative labeled with (14)C ([(14)C]PPD) with a biological potency equivalent to the International Standard for tuberculin PPD was used to study the retention of tuberculin PPD in the skin of sensitized and nonsensitized animals. We found that [(14)C]PPD was almost entirely cleared from the skin test site during the first 18 to 24 h after injection and that when approximately 5% of the initial concentration of [(14)C]PPD was present in the skin test site, the size of the tuberculin skin reaction in sensitized guinea pigs was at its maximum. Furthermore, the addition of 5 or 50 mug of Tween 80 per ml to a solution of PPD did not change either the rate of clearance of PPD from the skin test sites of sensitized guinea pigs or the size of the tuberculin skin reactions. There was no difference in the rate of clearance of [(14)C]PPD from the skin test sites between sensitized and nonsensitized guinea pigs and between guinea pigs of different age. However, there was a significant difference in the rate of clearance of [(14)C]PPD between the guinea pig and the mouse. Finally, the percentage of [(14)C]PPD retained in the site of injection at 24 h was in the neighborhood of 5% of the initial concentration of the solution of PPD injected. The significance of these phenomena is discussed.  相似文献   

8.
Immunobiologic activities of tuberculin preparations and their components have been comparatively studied using gel filtration and high-performance liquid chromatography (HPLC) techniques. It is shown that high-molecular weight fraction of protein-purified derivate tuberculin (PPD) had higher activity as compared to nonfractionated preparation in skin tests on guinea pigs sensitized with Mycobacterium bovis BCG as well as in enzyme-linked immunosorbent assay with affinity purified rabbit antibodies against PPD. Using the preparative HPLC-technique we failed to isolate a component of PPD having greater tuberculin test potency than nonfractionated preparation.  相似文献   

9.
Preparation, Purification, and Stability of Tuberculin   总被引:6,自引:1,他引:5       下载免费PDF全文
The method used to produce “Connaught” tuberculin purified protein derivative (PPD) is described. The tuberculin PPD for the multiple-puncture method was shown to be stable for at least 24 months at 5 C; tuberculin PPD for the intracutaneous method was shown to be stable at 5 C and 24 C for a period of 18 months in the presence of Tween 80. Evans blue or brillant vital red was added to tuberculin PPD for improved testing by the multiple-puncture method. These tinted tuberculin preparations were found to be as stable as the Connaught tuberculin PPD preparations without dye at 5 C. Freeze-dried tuberculin PPD with Plasdone as an inert base was found to be remarkably stable for a period of at least 24 months at 5, 24, and 37 C.  相似文献   

10.
Peritoneal exudate cells (PEC) and peripheral blood leukocytes (PBL) are most frequently used in the migration inhibition test. The aim o this work was to compare the ability of these two types of cells to reflect tuberculin hypersensitivity in the migration inhibition test. We sensitized 36 guinea pigs with complete Freund's adjuvant and 20 controls were injected with incomplete Freund's adjuvant. Migration of PEC in medium containing 5, 15, or 75 μg of PPD/ml was assessed after 30 min, and 1, 2, 4, 18, 24, and 48 hr of incubation. The migration of PEC from sensitized animals was inhibited, the inhibition being dose dependent and, with lower concentrations of the antigen, becoming significant only after 4 hr or later. With both PEC and PBL from the same sensitized animal we observed virtually identical migration inhibition in the presence of 75 μg of PPD/ml. A correlation was found between the migration inhibition indices of PEC and PBL. In the indirect test, active supernatants containing lymphokines caused nearly identical migration inhibition of PEC and PBL from normal animals. It follows that in the guinea pig PEC and PBL behave alike both in the direct and in the indirect migration inhibition tests. Thus, PEC and PBL appear to be equally valuable sources of cells for migration inhibition tests.  相似文献   

11.
Preparation of 14C-labeled tuberculin purified protein derivative   总被引:1,自引:1,他引:0       下载免费PDF全文
(14)C-labeled tuberculin purified protein derivative ((14)C-tuberculin PPD) has been prepared from culture filtrates of Mycobacterium tuberculosis var. hominis grown in a culture medium containing uniformly labeled (14)C-amino acids. With a mixture of (14)C-amino acids (an acid hydrolysate of (14)C-Chlorella protein) in the medium, the recoveries of (14)C in the final product were higher than with (14)C-labeled-l-glutamic acid. (14)C-tuberculin PPD was separated into tuberculoprotein and nucleic acid by paper electrophoresis. The specific radioactivity of tuberculoprotein was substantially greater than that of the nucleic acid. (14)C-tuberculin PPD is advocated as a means to measure the adsorption of tuberculin to glass or other surfaces. It could also prove useful as a means to study the structure and mode of action of tuberculin.  相似文献   

12.
The indirect agarose technique of leukocyte migration inhibition has been used to measure the response of human peripheral blood lymphocytes to several viruses. Using commercially available viral antigens, the indirect assay was found to be more sensitive than the direct agarose technique. Supernatants from cultures of sensitive lymphocytes with virus contained a nondialysable factor which inhibited the migration of polymorphonuclear leukocytes (PMN). Under strict conditions of assay, whereby all culture supernatants were tested together on the same PMN preparation, the degree of migration inhibition obtained in response to mumps virus correlated well with the size of the skin test reaction to mumps. A similar relationship was shown for PPD. A good correlation existed also between the degree of migration inhibition and the lymphocyte transformation response for each of these two antigens.  相似文献   

13.
Tuberculin purified protein derivative (PPD) has been prepared by seven different precipitation methods from culture filtrate of Mycobacterium tuberculosis var. hominis. It was found to contain 48 to 99% tuberculoprotein, depending on the method of precipitation. The remaining percentage is represented by nucleic acid, polysaccharide, and ash. Activation analysis on tuberculin PPD and on tubercle bacilli has revealed the presence of trace elements. The molecular weight of tuberculin PPD has been found to be of the order of 14,800 to 27,800. The biological activity of tuberculin PPD varies from lot to lot and from method to method. A correlation between its molecular weight and its biological activity seems to exist.  相似文献   

14.
Skin testing with 5 tuberculin units (TU) of purified protein derivative (PPD) of tuberculin stabilized with polysorbate (Tween) 80 was done 3 months and 1 year after immunization with bacille Calmette-Guérin (BCG) vaccine in two groups of children: one group vaccinated at birth and another group at age 6 years. Interpretation of the PPD skin test with 5 TU is possible in children 1 year and older vaccinated with BCG at birth: if the diameter of induration is more than 10 to 12 mm the reaction cannot be ascribed to BCG vaccination and is highly suggestive of supervening infection with Mycobacterium tuberculosis or occasionally atypical mycobacteria. In contrast, the interpretation of a PPD test in children vaccinated at age 6 years is extremely difficult.  相似文献   

15.
Human mononuclear leukocytes generate cell-bound procoagulant activity (LPCA) after incubation with an antigen (mumps or tuberculin) to which the donor was previously sensitized. An inhibitor of coagulation appears to be liberated into the extracellular culture fluid during incubation of leukocytes with the sensitizing antigen. Removal of this activity before measuring LPCA resulted in a reliable test that correlated directly with delayed skin reactivity. The assay was particularly sensitive in that cells from weakly sensitized donors who reacted only to high doses of tuberculin (100 TU) in the delayed skin tests produced detectable LPCA in vitro. By contrast cells from weakly sensitized donors did not react to PPD in the lymphocyte blast transformation test or the direct macrophage migration inhibition factor test. The LPCA assay correlated closely with the blast transformation and MIF tests in which cells were used from more strongly sensitized donors who reacted in skin tests with lower doses of tuberculin (1 or 10 TU). The assays were antigen-specific in that cells from donors sensitive to mumps antigen but not to tuberculin reacted only to mumps antigen in vitro. The assay was extremely reproducible; cells from individual donors reacted to the same extent over a period of 8 mo). We propose that the assay system reported here represents an improved method for the measurement of cell-mediated immunity in vitro because it requires fewer donor cells, is technically simpler, and is more sensitive than previously described methods.  相似文献   

16.

Background  

Bovine tuberculosis, caused by Mycobacterium bovis, afflicts approximately 50 million cattle worldwide and is detected by the tuberculin skin test (TST). While it has long been recognized that purified protein derivative (PPD) tuberculin is composed of a mixture of M. bovis derived protein components, little is known about the quality, relative quantity and identity of the proteins that make up PPD tuberculin. We manufactured a sterile filtered PPD tuberculin (SF-PPD) from a nine-week-old M. bovis culture supernatant in order to characterise the culture filtrate proteins (CFP) which make up M. bovis PPD tuberculin and to compare the antibody response of M. bovis infected versus M. bovis sensitized cattle.  相似文献   

17.
The antibody responses to the recombinant analogs of Mycobacterium bovis antigens rMPB63 and rMPB83, as well as to tuberculin PPD, were determined in 96 bovine blood serum specimens. A statistical approach based on the approximation with multiple Gaussian curves with Levenberg-Marquardt algorithm was proposed for analysis of the distribution of antibody levels to the studied antigens. The results demonstrate that the recombinant MPB63 and MPB83 antigens along with the traditionally used PPD are appropriate for designing the test kits for diagnosing bovine tuberculosis at a herd level.  相似文献   

18.
Cyclophosphamide in a single dose of 300 mg per kg, injected intraperitoneally 3 days before the sensitization of guinea pigs with azobenzenearsonate-N-acetyl-1-tryosine in complete Freund's adjuvant, suppressed the development of delayed hypersensitivity to PPD tuberculin and Ars-insulin. Peritoneal cell migration inhibition induced in vitro by PPD or Ars-Tyr was also suppressed by the Cy pretreatment. It was concluded that Cy suppresses cell-mediated immunity directly, i.e., in a system, in which no B cell response is available as a target for Cy (the response to ArsTyr).  相似文献   

19.

Background

Purified protein derivative (PPD) has been used for more than half a century as an antigen for the diagnosis of tuberculosis infection based on delayed type hypersensitivity. Although designated as “purified,” in reality, the composition of PPD is highly complex and remains ill-defined. In this report, high resolution mass spectrometry was applied to understand the complexity of its constituent components. A comparative proteomic analysis of various PPD preparations and their functional characterization is likely to help in short-listing the relevant antigens required to prepare a less complex and more potent reagent for diagnostic purposes.

Results

Proteomic analysis of Connaught Tuberculin 68 (PPD-CT68), a tuberculin preparation generated from M. tuberculosis, was carried out in this study. PPD-CT68 is the protein component of a commercially available tuberculin preparation, Tubersol, which is used for tuberculin skin testing. Using a high resolution LTQ-Orbitrap Velos mass spectrometer, we identified 265 different proteins. The identified proteins were compared with those identified from PPD M. bovis, PPD M. avium and PPD-S2 from previous mass spectrometry-based studies. In all, 142 proteins were found to be shared between PPD-CT68 and PPD-S2 preparations. Out of the 354 proteins from M. tuberculosis–derived PPDs (i.e. proteins in either PPD-CT68 or PPD-S2), 37 proteins were found to be shared with M. avium PPD and 80 were shared with M. bovis PPD. Alignment of PPD-CT68 proteins with proteins encoded by 24 lung infecting bacteria revealed a number of similar proteins (206 bacterial proteins shared epitopes with 47 PPD-CT68 proteins), which could potentially be involved in causing cross-reactivity. The data have been deposited to the ProteomeXchange with identifier PXD000377.

Conclusions

Proteomic and bioinformatics analysis of different PPD preparations revealed commonly and differentially represented proteins. This information could help in delineating the relevant antigens represented in various PPDs, which could further lead to development of a lesser complex and better defined skin test antigen with a higher specificity and sensitivity.  相似文献   

20.
Transfer factor (TF) causes nonimmune lymphocytes to produce leukocyte migration inhibitory factor (LMIF) in the presence of purified protein derivative (PPD). The activity of TF was measured by leukocyte migration inhibitory test (LMIT). The LMIT was a modification of the conventional agarose droplet method. To express the activity of LMIF quantitatively and simply, LMIF titer was introduced. The LMIF titer was obtained from the combination of two factors, LMIF dilution and cell migration diameter, and therefore this made the LMIT much more sensitive as compared to the conventional LMIT. The responsiveness of lymphocytes from acute lymphoblastic leukemia (ALL) and from cell-mediated immunodeficiency in children to TF was assayed by LMIT. In ALL, the lymphocyte responsiveness was poor in relapse but improved with remission. The responsiveness was remarkably well in 3 patients with cell-mediated immunodeficiency. This method appears useful for the in vitro evaluation of responsiveness of lymphocytes to TF.  相似文献   

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