Role of serum and pleural adenosine deaminase activity compared to pleural fluid analysis in patients with pleural effusion of various etiology

Mubasheer Ali; Prashant Janjal; MV Raghavendra Rao

doi:10.4103/mjhs.mjhs_27_22

ORIGINAL ARTICLE

Year : 2023 | Volume : 11 | Issue : 1 | Page : 34-40

Role of serum and pleural adenosine deaminase activity compared to pleural fluid analysis in patients with pleural effusion of various etiology

Mubasheer Ali¹, Prashant Janjal², MV Raghavendra Rao³
¹ Department of General Medicine, Shadan Institute of Medical Sciences, Hyderabad, Telangana, India
² Public Health Specialist, Apollo Telehealth, Apollo Institute of Medical Sciences and Research, Hyderabad, Telangana, India
³ Department of General Medicine, Apollo Institute of Medical Sciences and Research, Hyderabad, Telangana, India

Date of Submission	27-Jun-2022
Date of Decision	21-Aug-2022
Date of Acceptance	29-Aug-2022
Date of Web Publication	02-Feb-2023

Correspondence Address:
Mubasheer Ali
Department of General Medicine, Shadan Institute of Medical Sciences, Hyderabad, Telangana
India

Source of Support: None, Conflict of Interest: None

DOI: 10.4103/mjhs.mjhs_27_22

Abstract

Background: Numerous tests are available for determining the cause of exudate, but all these tests lack sensitivity and specificity, and they are not generally available. Adenosine deaminase (ADA) activity in pleural or peritoneal fluid is simple and useful investigation in diagnosis of exudate effusion, particularly in differentiating tuberculous from nontuberculous exudative effusion.
Objectives: To estimate the diagnostic utility of pleural and peritoneal fluid ADA/serum ADA ratio, in the diagnosis of tubercular pleural effusion.
Materials and Methods: A hospital-based cross-sectional diagnostic evaluation study was carried out among 50 patients with pleural effusions selected to assay ADA activity in pleural fluid and serum along with pleural fluid analysis. Effusions were classified into transudative and exudative after careful evaluation of all biochemical parameters of pleural fluid. Cutoff value for pleural ADA was taken as 60 U/L and for pleural/serum ADA was taken as 2.5.
Results: Forty-three patients had exudative effusions among which 38 were tubercular and five were nontubercular, seven cases were transudative. Mean pleural ADA levels in tuberculous group (80.31 ± 24.84 U/L) were higher (P < 0.01) than nontubercular group (23.00 ± 5.22 U/L). Serum ADA levels in tubercular group (27.23 ± 7.32 U/L) were significantly higher (P < 0.0l) as compared to nontubercular group (14.95 ± 7.04 U/L). Serum ADA levels in tubercular group (27.23 ± 7.32 U/L) were also significantly higher as compared to the control group of healthy individuals (13.00 ± 2.75 U/L). Pleural ADA cutoff at 60 U/L gave a sensitivity and specificity of 81% and 100%, respectively, whereas pleural serum ADA ratio at a cutoff of 2.5 gave a sensitivity and specificity of 71% and 81%, respectively.
Conclusion: The measurement of ADA in tubercular pleural effusion is not only relevant, but also of a high diagnostic value when other clinical and laboratory tests are either negative or inconclusive.

Keywords: Adenosine deaminase, pleural adenosine deaminase, serum adenosine deaminase, tubercular pleural effusion

How to cite this article:
Ali M, Janjal P, Raghavendra Rao M V. Role of serum and pleural adenosine deaminase activity compared to pleural fluid analysis in patients with pleural effusion of various etiology. MRIMS J Health Sci 2023;11:34-40

How to cite this URL:
Ali M, Janjal P, Raghavendra Rao M V. Role of serum and pleural adenosine deaminase activity compared to pleural fluid analysis in patients with pleural effusion of various etiology. MRIMS J Health Sci [serial online] 2023 [cited 2023 Mar 30];11:34-40. Available from: http://www.mrimsjournal.com/text.asp?2023/11/1/34/369040

Introduction

Pleural effusion and ascites are the common problems in clinical practice. These effusions may transudative or exudative. If the effusion is a transudate, no further diagnostic procedure is required, and in the majority of cases, the underlying cause can be diagnosed clinically, for example, congestive heart failure, hypo-proteinemia, cirrhosis of liver, etc. However, if the effusion is an exudate, more investigation is needed to define the cause of effusion.^[1]

Numerous tests are available for determining the cause of exudate, but all these tests lack sensitivity and specificity, and the tests are not generally available. In light of this observation, adenosine deaminase (ADA) activity in pleural or peritoneal fluid has been found to be simple and useful investigation in the diagnosis of exudate effusion, particularly in differentiating tuberculous from nontuberculous exudative effusion. Excessive and abnormal accumulation of fluid in the pleural cavity is defined as pleural effusion. Pleural effusion is a common complication of many disease processes. Tuberculosis (TB) is still one of the most important causes of exudative pleural effusion. TB pleural effusion occurs in approximately 5% of patients with Mycobacterium TB infection. The HIV pandemic has been associated with a doubling of the incidence of extra pulmonary TB, which has resulted in increased recognition of TB pleural effusions even in developed nations.^[2]

A definite diagnosis of tuberculosis pleural effusion (TPE) can be difficult to make because of the low sensitivity and/or specificity of noninvasive traditional diagnostic tools. In most series of patients with TPE, the results of pleural fluid staining for acid-fast bacilli are virtually always negative, and pleural fluid cultures are positive for mycobacterium in <25% of cases. On the other hand, a pleural biopsy specimen will demonstrate granulomatous pleuritis in 80% of patients with TPE, and when a culture of a biopsy specimen is combined with histological examination, the diagnosis can be established in approximately 90% of cases.^[3]

TB pleurisy is thought to be the result of a delayed hypersensitivity reaction in response to the presence of mycobacterial antigens in the pleural space.^[4] This immunologic reaction causes the stimulation and differentiation of lymphocytes, which release lymphokines, which in turn activate macrophages for an enhanced bactericidal effect.^[4] The levels of ADA, an enzyme found in most cells, are increased in TB pleural effusions, and this determination has acquired popularity as a diagnostic test in high-incidence areas for TPE because it is not invasive, the assay is not expensive, and it is readily accessible. ADA activity has been proposed as a diagnostic test for tubercular pleurisy since 1985. Many workers Ocaña et al. 19836, Tom Patterson 1984, others^[5],[6] from western countries and Sinha et al. 1985^[7] and others^[8],[9] from India in a number of studies have shown pleural ADA activity in TB effusion is raised compared to malignant and para pneumonic effusions. ADA is involved in the proliferation and differentiation of lymphocytes, especially T lymphocytes. They release ADA when stimulated in the presence of live intracellular microorganisms.^[3] For this reason, ADA has been looked on as a marker of cell-mediated immunity and, in particular, as a marker of the activation of T lymphocytes. Several reports have suggested that an elevated pleural fluid ADA level predicts tuberculous pleuritis with a sensitivity of 90%–100% and a specificity of 89%–100%.^[8]

With this background, present study was carried out to estimate ADA levels in pleural and peritoneal fluid to distinguish between Tubercular and non-Tubercular effusion and to estimate the diagnostic utility of pleural and peritoneal fluid ADA/serum ADA ratio, in the diagnosis of tubercular pleural effusion

Materials and Methods

Institutional Ethics Committee permission was obtained. Written informed consent was taken from all eligible participants. Present cross-sectional diagnostic evaluation study was conducted at Princess Esra Hospital and Owaisi Hospital and Research centre, Deccan College of Medical Sciences. The study group consisted of 50 cases of pleural effusion and 25 cases of ascites who got admitted in the hospital during the study period, including both sexes.

Each patient was carefully evaluated based on history, clinical examination and investigations. The diagnosis was established by detailed history, clinical examination, blood investigations, chest X-rays, ultrasonography abdomen and pleural and peritoneal fluid analysis. In all the patients' serum ADA and pleural/peritoneal fluid ADA values were estimated, serum ADA in 25 healthy volunteers was estimated as control group. The cut off value of pleural fluid ADA was kept at 60 U/L and peritoneal fluid ADA was kept at 40 U/L. Serous fluid/serum ADA ratio was also calculated and its utility was assessed. The cut off value of pleural/peritoneal fluid/serum ADA ratio was taken as 2.5. To meet our objectives, various parameters have been taken up like comparing pleural and peritoneal fluid ADA between tuberculous and nontuberculous group, serum ADA between tuberculous and nontuberculous group and also between tuberculous and control groups. And lastly serous/serum ADA ratio has been taken into account and compared between the tuberculous and nontuberculous groups.

Patients with confirmed diagnosis of pleural effusion based on history, clinical features, pleural effusion symptoms and signs later verified by radiology were included in the present study. Patients diagnosed with empyema, chylothorax, lung malignancy/abdominal malignancy/secondary metastasis, Rheumatoid arthritis and sarcoidosis, immuno-compromised states were excluded.

All patients underwent investigations like Hemoglobin, white blood cell count: Total and differential count, ESR, Urea expressed in gm%, Serum creatinine Expressed in gm%, Urine examination: Albumin, sugar and Microscopy, Sputum for AFB: Done by ZN method, Plain X-ray chest: P/A view and lateral view, Ultrasound of abdomen, Mantoux test, Pleural/peritoneal fluid analysis: Colour Specific gravity by urinorneter Protein by Bioret method. All the investigations were done as per the standard guidelines and procedures. Estimation of ADA in Pleural/Peritoneal Effusion was done using Guisti and Galanti method.

Data was entered in the Microsoft Excel worksheet and analysed using proportions, mean values. Difference in proportions was tested by Chi-square and difference in mean was tested by t-test for two groups and ANOVA for more than two groups. At cut off of 0.05 or less, P value was taken as statistically significant. For assessing the diagnostic utility, sensitivity and specificity was calculated.

Results

In our study a total of 50 cases of pleural effusions and were 25 cases of peritoneal effusions were studied and following observations were made.

Majority cases (76%) had tuberculous pleural effusion, whereas in peritoneal effusion, transudative type was most common in 32% [Table 1].

Table 1: Distribution of study subjects as per different variables

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Most common symptom of pleural effusion was fever (88%) followed by cough (82%) while it was loss of appetite and distension of abdomen (96%) followed by pain abdomen (80%) in peritoneal effusion. Most common respiratory sign in pleural effusion was stony dullness (100%) followed by decreased VF. VR (98%). Most common sign in peritoneal effusion was shifting dullness (100%) followed by icterus (48%) [Table 2].

Table 2: Distribution of the presenting symptoms and signs of pleural and peritoneal effusion

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The mean ESR at the end of first hour was significantly more in tuberculous pleural effusion compared to synpneumonic and transudative types of pleural effusion (P < 0.05). Similarly, in case of peritoneal effusion also the mean ESR at the end of first hour was significantly more in tuberculous peritoneal effusion compared to bacterial and transudative types of pleural effusion (P < 0.05). The pleural fluid protein was significantly less in transudative type of pleural effusion compared to tuberculous and Synpneumonic types of pleural effusion.(P < 0.05) [Table 3].

Table 3: Comparison of erythrocyte sedimentation rate values and pleural fluid protein in various types of pleural effusion

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The mean pleural ADA level was significantly more in tuberculous type of pleural effusion compared to synpneumonic and transudative types of pleural effusion (P < 0.05). The mean peritoneal ADA level was significantly more in tuberculous type of peritoneal effusion compared to bacterial and transudative types of peritoneal effusion (P < 0.05) [Table 4].

Table 4: Comparison of mean adenosine deaminase level of pleural/peritoneal fluid in different types of effusion

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The pleural ADA levels were significantly raised in tubercular type of pleural effusion compared to non-tubercular type (P < 0.05). The peritoneal ADA levels were significantly raised in tubercular type of peritoneal effusion compared to non-tubercular type (P < 0.05) [Table 5].

Table 5: Comparison of pleural/peritoneal adenosine deaminase level for tubercular and nontubercular types

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The mean serum ADA levels were significantly raised in tuberculous type of pleural effusion compared to synpneumonic and transudative types of pleural effusion (P < 0.05). The mean serum ADA level was significantly more in tuberculous type of peritoneal effusion compared to bacterial and transudative types of peritoneal effusion (P < 0.05) [Table 6].

Table 6: Comparison of mean serum adenosine deaminase level of pleural/peritoneal fluid in different types of effusion

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When compared to controls, the mean ADA levels in Tubercular pleural effusion as well as in Tubercular peritoneal effusion were significantly more (P < 0.05) [Table 7].

Table 7: Comparison of mean serum adenosine deaminase levels among effusion and controls for tubercular and nontubercular types

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Mean pleural fluid ADA/serum ADA ratio was significantly more in tuberculous type of pleural effusion compared to Synpneumonic and transudative types of pleural effusion (P < 0.05). The mean pleural fluid ADA/serum ADA ratio was significantly more in tuberculous type of peritoneal effusion compared to bacterial and transudative types of peritoneal effusion (P < 0.05) [Table 8].

Table 8: Distribution of mean pleural fluid adenosine deaminase/serum adenosine deaminase ratio in different types of effusion

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Diagnostic utility of pleural ADA in tubercular pleural effusion at cut point of 60.0 U/L was very good with sensitivity of 81% and 100% of specificity. The sensitivity and specificity of pleural ADA/serum ADA ratio in tubercular peritoneal effusion was 86% and 100% respectively at cut point of 40.0 U/L. the sensitivity and specificity of pleural serum ADA ratio in tubercular pleural effusion was 71% and 81% respectively at cut point of 2.5. The sensitivity and specificity of pleural serum ADA ratio in tubercular pleural effusion respectively at cut point of 2.5 [Table 9].

Table 9: The utility of adenosine deaminase

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Discussion

The age of the patients in this study ranged from 17 to 70 years (mean 43.76 ± 15.25 for pleural effusion and 41 ± 16.91 for peritoneal effusion). This is consistent with the series published by McAleer et al.^[9] where the age ranged from 14 to 85 years and with Ocaña et al.^[5] where age ranged from 8 to 85 years with an average age of 45.

Out of 38 tuberculous patients 25 (65%) and 13 (35%) were females. In McAleer et al.^[9] series of 118 cases 63 cases (53%) were males and 55 (47%) were females. Levine et al.^[10] found an incidence of 65% in males and in 35% females in his series. The findings of this study are almost consistent with previous studies.

Most common symptom of pleural effusion was fever (88%) followed by cough (82%) while it was loss of appetite and distension of abdomen (96%) followed by pain abdomen (80%) in peritoneal effusion. Most common respiratory sign in pleural effusion was stony dullness (100%) followed by decreased VF. VR (98%). Most common sign in peritoneal effusion was shifting dullness (100%) followed by icterus (48%).

In present study the pleural fluid ADA levels were raised in all cases. The ADA levels were compared between tuberculous and nontuberculous groups and the mean ADA in tubercular effusion (80.31 ± 24.84) was significantly higher (P < 0.00l) as compared to other causes of effusion (synpneumonic - 24.0 ± 5.19 and transudative (22.28 ± 5.52) and it is helpful in the differential diagnosis of pleural effusion of various etiology. Earlier studies by Smach et al.^[11] and Neves et al.^[12] have shown similar results. The present study correlates well with the above studies. Ocaña et al.^[5] has reported using a cut-off value of 50 U/L, ADA has shown a sensitivity of 100% and specificity of 87%. In our study using the cutoff value of 60U/L for tuberculous group, ADA showed sensitivity and specificity of 81.5% and 100% respectively. It was determined that a pleural fluid ADA level >70 IU/L is highly suggestive of TB, while a level <40 lU/L virtually excludes the diagnosis.^[13] A Meta-analysis^[14] of 40 studies published from 1966 to 1999 concluded that the test performance of ADA (sensitivity range 47.1%–100%, and specificity range 0%–100%) in diagnosing TB pleural effusions is reasonably good (adequate to avoid pleural biopsy in young patients from areas with high prevalence of TB).

Chopra and Singh^[15] have found a direct correlation between pleural fluid ADA and T-lymphocytes percentage in TB patients. Pettersson et al.^[16] suggested that higher ADA activity in pleural fluid was probably due to ADA being synthesized by cells in the pleural cavity in tuberculous effusion; a reflection of local activity of selectively sequestrated T lymphocytes. In present study, higher lymphocyte predominance was noted in tuberculous pleural effusion, recently two isoenzymes have been identified namely ADA-l and ADA-2. ADA-1 isoenzymes have been found to be present in all the cells with highest activity observed in lymphocytes and monocytes. ADA-1 isozyme patient with Tubercular pleural effusion had increased ADA activity and ADA-2 isozyme to be primarily responsible for total activity. It was found that the determination of isozyme would enhance the diagnostic utility of ADA activity in the determination of pleural effusion. The cutoff value of the pleural fluid to serum ADA ratio in our study in case of tubercular pleural effusion was kept more than 2.5. This value showed a sensitivity and specificity of 71% and 81% respectively.

Patterson et al.^[17] in his study calculated the tubercular pleural ADA and serum ADA ratio and found that the value above 2.5 always denote tubercular pleural effusion. In present study ratio is a171, p-ADA/s-ADA ratio was used for diagnosis of tuberculous pleural effusion, a threshold value of 1.8 gave sensitivity and specificity of 82.6% and 84.8% respectively. In another study by Jadhav and Bardapurkar^[18] at a cut-off point 1.28, pleural fluid/serum ADA ratio was found to have sensitivity 84% and specificity 90%, respectively. From this study it is concluded that, ADA is a useful biochemical marker to suggest exudative effusions. Mean serum ADA values were calculated in all the 50 cases of pleural effusion and also in 25 healthy controls. Mean serum ADA concentration in healthy controls in our study was 13 ± 2.75 U/L. In earlier studies done by Jhamaria and Jenaw^[19] and Nagaraja et al.^[20] mean serum ADA concentration in healthy controls were 19.09 U/L and 14.56 U/L respectively. Our study correlates well with the above studies. Jhamaria and Jenaw^[19] shows the serum ADA levels in tubercular pleural effusion in their respective studies were 42.44 U/L and 23.38 U/L respectively which was significantly elevated compared to other causes of pleural effusion. In our study the mean serum ADA in study group showed elevated ADA levels in tubercular effusion (27.23 ± 732) compared to synpneumonic effusion (11.2 ± 4.9) and transudative effusion (18.7 ± 9.17) which is statistically significant (P < 0.05) indicating similarity with the above studies.

In the present study, the peritoneal fluid ADA levels were raised in all cases of tuberculous peritonitis. The mean ADA levels in tuberculous peritoneal effusion (56 ± 21.26U/L) were significantly higher (P < 0.001) as compared to the other causes (bacterial-12 ± 2.82 U/L and transudative -16.43 ± 7.32 U/L) and is helpful in differential diagnosis of peritoneal effusions of various etiology. Earlier studies by Ocaña et al.,^[21] and Bhargava et al.^[22] all demonstrated an increased ADA activity in the ascites of TB patients because of high prevalence of TB and also they had a large study group.

Mean serum ADA values were calculated in all 25 cases of peritoneal effusions and also in 25 healthy controls. Mean serum ADA concentrations in healthy controls in present study was 13 ± 2.74 U/L. In present study the mean serum ADA levels in study group showed elevated ADA levels in tubercular peritonitis (24.14 ± 17.18 U/L) compared to bacterial peritonitis (12 ± 2.82 U/L) and transudative effusions (16.43 ± 7.32 U/L) which is statistically significant (P < 0.001). Pleural fluid ADA value of tuberculous group in the present study is correlating with the other studies done in India, like the study done by Gupta et al.^[23] and Sinha et al.^[7] where the values are slightly lower than the present study. In nontuberculous group also the values are correlating with the study done by Ocaria I et al.^[21]

Conclusion

Pleural fluid ADA estimation is a useful tool in the diagnosis of pleural effusion especially tubercular variety. ADA activity is definitely raised in cases of tubercular effusions as compared to non-tuberculous type. Serum ADA levels also may remain increased in cases of TB, but its value is inconclusive for taking any decision. A pleural ADA cut-off value of more than 60 U/L is virtually diagnostic of tuberculous pleural effusion. Pleural ADA/serum ADA ratio with a cutoff of 2.5 may be a useful tool to diagnose tubercular pleural effusion, but further studies with larger study population is required for a definitive inference. Estimation of pleural ADA alone can help us differentiate transudative and exudative pleural effusions. ADA is a simple, reliable, cheaper diagnostic tool at our hands as compared to pleural biopsy which is an invasive technique with complications and also pleural fluid culture which takes weeks together for mycobacteria to be isolated.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

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