This clinical study was conducted to evaluate the diagnostic value of C-reactive protein (CRP), interlekin-6 (IL-6) and immunoglobulin M (IgM) in the early diagnosis of neonatal sepsis. The diagnostic values of each marker separately or in combinations were evaluated. The optimal cut-off values of each marker in the diagnosis of neonatal sepsis were defined. Between December 2004 and March 2005, a total of 78 neonates at different ages with different diagnoses in a neonatal intensive care unit in North Jordan were enrolled. Patients were classified into ‘sepsis’ group, ‘probable sepsis’ group and ‘no sepsis’ group. Blood samples were collected for CRP, IL-6 and IgM determination. A CRP value of 5 mg/l was the best among the three parameters with 95% sensitivity and 98% negative predictive value. Combination between parameters was helpful in enhancing the ability to diagnose sepsis. The best combination was CRP ≥ 5 mg/l and/or IgM of ≥20 mg/dl. We conclude that CRP, IL-6 and IgM are helpful in the early diagnosis of Gram-negative neonatal sepsis. However, CRP continues to be the best single test. The use of both CRP and IgM in combination was the most helpful in predicting Gram-negative neonatal sepsis. We speculate a significant role of this combination in making decisions regarding antibiotics treatment and upgrading the level of medical care and observation in a setting where Gram-negative micro-organisms are causing the majority of neonatal infections.