The aim of this study was to investigate the relationships among technetium-99m tetrofosmin (Tc-TF) accumulation in untreated small cell lung cancer (SCLC), the expression of P-glycoprotein (Pgp) and multidrug resistance related protein-1 (MRP1), and the response to chemotherapy in patients with untreated SCLC. Thirty patients with SCLC were studied with chest scintigraphy 15 to 30 min after intravenous injection of Tc-TF before chemotherapeutic induction. Tc-TF chest scans were interpreted both visually and quantitatively. The response to chemotherapy was evaluated upon completion of chemotherapy. Immunohistochemical analyses were performed on multiple non-consecutive sections of biopsy specimens to detect Pgp and MRP1 expression. Fifteen patients with good response to chemotherapy had a significantly higher incidence (100.0%) of positive Tc-TF chest single photon emission computed tomography (SPECT) findings and negative Pgp or MPR expression than 15 patients with poor response (20%) (P<0.05). The tumor/background (T/B) ratios were 1.8+/-0.3 and 1.2+/-0.3 for patients with good response and poor response, respectively (P<0.05). However, other prognostic factors (performance status, tumor size and stage) were not significantly related to Tc-TF chest scan findings and response to chemotherapy. Tc-TF chest scintigraphy correlated well with Pgp or MRP1 expression and accurately predicted the response to chemotherapy in patients with SCLC.