Comparative characteristics of immunohistochemical detection of M. tuberculosis antigens and mycobacteria by Ziehl–Neelsen method in lung tissue with tuberculoma during progressive course of tuberculosis

Abstract

The aim of the study – to compare the character of expression of M. tuberculosis antigens in immunohistochemical (IHC) study and detection of acid-fast bacilli (AFB) by Ziehl–Neelsen method in lung tissue with tuberculoma in the chronic course of tuberculosis during the exacerbation phase.Materials and methods. The materials for the study were resected lungs, in total 19 cases. The thickness of the serial sections was 5–6 microns. Staining was carried out with hematoxylin and eosin, by Ziehl–Neelsen, and the IHC.IHC study used a rabbit polyclonal antibody to Mycobacterium tuberculosis. Antibody (USA) was used at 1:2500 dilution. Assessing of the positive reaction was according to the conditional scale: + – background color, pale yellow, diffuse; ++ – fine-granular (light brown); +++ – coarse-granular (intense brown). Quantitative gradation of AFB was the following: out or within macrophages in one field of view: few, 1–5 units; a moderate amount – 6–15, and many – more than 15.Results. In all the cases pulmonary tuberculosis had morphological signs of the progression of a specific inflammatory process.Ziehl–Neelsen stain showed that the most frequently free located AFB were detected in the necrotic nucleus (84.2 %) and granulation layer of the capsule of tuberculoma (68.4 %). Most often single bacilli were noted. A greater number of bacilli were found inside macrophages.The most frequently intracellular bacilli were found in macrophages located in alveolar spaces – 84.2 % observations in a moderate number.IHC study showed that in all the cases, cells with a coarse-granular positivity (+++) to MBT antigens were determined. Macrophages within the alveoli outside tuberculoma, with same positivity were in 94.7 %; fewer – macrophages and epithelioid cells were in the granulation layer of the tuberculoma capsule, a total of 84.2 %. The percentage of observations of a large number of cells in the alveoli reached 77.8 %, and in the granulation layer of the capsule – 30.0–50.0 %.More often the fine-granular staining of cells in the same parts of lung tissue was observed, a cases of large and/or moderate number of cells prevailed.Conclusions. The AFB was identified in 94.7 % cases by Ziehl–Neelsen method. In IHC study a positive reaction of macrophage cells was detected in all the cases (100 %).IHC study showed that maximum antigenic load was determined in macrophages localized inside alveoli outside of tuberculoma and slightly less in macrophage cells of the granulation layer of the tuberculoma’s capsule. The results of both methods for detecting mycobacteria and/or their fragments demonstrated the coincidence of the maximum number of observations of localization of the bacilli structures, namely in granulation layer of the tuberculoma’s capsule and the preserved alveolar spaces outside the tuberculoma.