BACKGROUND: Anti-neutrophil cytoplasm antibodies (ANCA) play an important role as specific and sensitive markers for small vessel vasculitis and in some other systemic disorders. Indirect immunofluorescence test, known as the "Gold Standard" for screening of ANCA, can be further substantiated by ELISA for confirmation and for identifying sub-specificities like anti-Myeloperoxidase (anti-MPO), anti-Proteinase 3 (anti-PR3) and anti-Lactoferrin (anti-LF). AIMS: The present study was undertaken to investigate the incidence, specificities and strength of ANCA in suspected vasculitis cases and to correlate their presence with that of these auto-antibodies and with the disease. SUBJECTS AND METHODS: Sera from 130 clinically suspected vasculitis patients were studied. Indirect immunofluorescence microscopy (IIF) was used to identify cytoplasmic (c-ANCA), perinuclear (p-ANCA) and atypical (X-ANCA) patterns using ethanol and formalin fixed polymorphonuclear cells (PMN) and HL-60 cells from a human promyelocytic leukaemic cell line as substrates. ELISA was performed for identifying ANCA sub-specificities to anti-MPO and anti-PR3 and HEp-2 cells were used for detection of anti-nuclear antibodies (ANA). RESULTS: ANCA positivity was noted in 42.3% of these patients, wherein p-ANCA positivity rate was 34.6% and c-ANCA positivity was noted in 5.4% subjects. Three patients showed the unusual X-ANCA positivity. ELISA determined the sub-specificities: Out of 45 p-ANCA positive patients, 38 patients (84.4%) had anti-MPO and out of 7 c-ANCA positive patients, 5 patients (71.4%) had anti-PR3 antibodies. One patient with Class IV Lupus Nephritis, showed both anti-MPO and anti-PR3 antibodies and 17.8% p-ANCA positive patients had anti-Lactoferrin antibodies. CONCLUSIONS: Use of the Immunofluorescence method coupled with identification of ANCA sub-specificities by ELISA, is recommended for detection of ANCA in clinically suspected cases of small vessel and other systemic vasculitis.