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The concentration of C-Reactive Protein (C-RP) and Tumor Necrosis Factor alpha (TNF-α) were determined in sera of thirty neonates aged 0-30 days with early onset and late onset septicemia between November 2006-August 2007 in Ile-Ife, Nigeria. Determination of the concentration of C-RP and TNF-α as indicators of sepsis for prompt diagnosis of the condition possibly to avert fatality and morbidity in these neonates. Blood samples were intravenously obtained for bacterial culture and ELISA technique of 30 neonates and 10 control subjects. The concentration of C-RP and TNF-α were determined using (ELISA) technique. Bacterial isolates were cultured on selective and differential media from serum samples initially introduced into thioglycolate broth. Samples that showed growth were further studied. Staphylococci were confirmed as S. aureus by coagulase production on slide and tube tests using pooled human plasma. Coagulase negative staphylococci were spceciated using carbohydrates. Gram negative rods were characterized using conventional media and antibiotic sensitivity tests of isolates were carried out by the disc diffusion method. The result showed 56.7% (17) of the neonates were categorized as early onset septicemia and 43.3% (13) with late onset condition. The mean C-RP concentration in the sera of neonates with early and late onset septicemia were 10.61±1.9 and 11.25±2.5, respectively compared to controls 2.29±0.89 each. While for TNF-α 38.7±14.9 and 33.01±16.9 for early and late onset sepsis compared to control was 6.8±1.4 pg L-1. The mean C-RP concentration in bacteremic neonates was 10.50±1.9 mg L-1 compared with control 2.29±0.89. However, TNF-α mean serum concentration in same subjects was 38.04±14.19 pg L-1 compared with 6.8±1.4 pg L-1 for g positive organisms. While for gram negative organism the result was 10.55±1.94 mg L-1 against 2.29±1.94 mg L-1 for control. TNF-α value was 40.68±17.19 for g negative culture and 6.8±1.4 for control. Gram negative rods predominated (56.4%) with E. coli being 10% of total g negative rods. Coagulase staphylococci were the single most common isolates recovered. S. capitis 12.8% was the predominant staphylococci followed by S. aureus 7.7% and S. saprophiticus 2.6%. While C-RP and TNF-α are useful markers in early diagnosis of septicemia, elevation in C-RP concentrations may not necessarily indicate infection. With regards to TNF-α, elevation of this protein tend to correlate with infection. The study suggests both molecules are useful for evaluating early sepsis. Antibiotics sensitivity tests showed the bacterial isolates were susceptible mostly to quinolones while cotrimoxazole was the least effective.