Kupert et al. BMC Cancer 2011, 11:513 http://www.biomedcentral.com/1471-2407/11/513 Statistical Analysis The means and standard deviations were calculated with confirmed significant difference in plasma sPLA2-IIa level between lung cancer specimens and benign nodule specimens. Geometric means and standard deviations were also calculated with confirmed significant differ- ence in plasma sPLA2-IIa level between lung cancer specimens and clinical control specimens. Unpaired t test with Welch correction was performed to evaluate the difference between sPLA2-IIa means of (i) 96 lung cancer specimens versus 29 benign SPN specimens from the BNLCC; (ii) 44 T1 stage lung cancer specimens ver- sus 18 T2 stage lung cancer specimens from the BNLCC; (iii) 44 lung cancer specimens from the GELCC versus 29 benign SPN specimens from the BNLCC. Unpaired t test with Welch correction was also used to analyze overall cancer survival year associated with plasma sPLA2-IIa level from the BNLCC. A parametric Receiver Operating Characteristic (ROC) analysis of plasma sPLA2-IIa values was performed to evaluate the ability of the levels of plasma sPLA2-IIa to distinguish 96 patients with lung cancer from 29 patients with benign SPNs from the BNLCC. 44 lung cancer specimens from the GELCC relative to 29 benign SPNs specimens from the BNLCC were also subjected to the ROC analysis. The optimum cutoff value of plasma sPLA2-IIa was determined by calculating the Youden Index [25], which separated the combined set of sPLA2-IIa values into two groups, such that the number of correctly classified specimens was maximized, and the associated sensitivity and specificity for predicting lung cancer versus non-malignant nodule were determined. Results The levels of plasma sPLA2-IIa are increased in lung cancer patients The means and standard deviations of plasma sPLA2-IIa levels from 96 patients with lung cancer and 29 patients with benign lung nodules (SPNs) from the BNLCC were 3646 ± 407.3 and 1772 ± 306.8 pg/ml, respectively (Table 1 and 2, and see Additional file 1). Based on an unpaired t-test, the average plasma sPLA2-IIa level in lung cancer patients was significantly higher than that in the non-malignancy SPN controls (P = 0.004). The opti- mum cutoff value of plasma sPLA2-IIa by ROC analysis was 2.4 ng/ml, which resulted in 48% sensitivity and 86% specificity for predicting the presence of lung can- cer (Figure 1a and Table 3). The area under the curve (AUC) was 0.68 (95% CI: 0.58 ~ 0.79) (Figure 2b). We also analyzed 44 lung cancer samples from the GELCC in which mean and standard deviation of plasma sPLA2-IIa were 3718 ± 407.8 (see Additional file 2). The average plasma sPLA2-IIa level in these lung cancer patients was also significantly higher than that in Table 1 Characteristics of the patients BNLCC-Lung cancer Mean age year (range) M/F NSCLC SCLC Stage Blooddraw before resection Blooddraw after resection BNLCC-benign SPN M/F M/F Mean age year (range) Blooddraw before resection Blooddraw after resection GELCC-Lung cancer Mean age year (range) Healthy donor Mean age year (range) Adenocarcinoma Squamous carcinoma Other NSCLC I II III+IV Page 3 of 10 96 37/59 64 (41~88) 93 54 21 18 3 44 18 4 96 0 29 29 0 44 20 15/14 56 (31~81) 18/26 61 (47~70) 49 (24~65) patients with benign SPNs from the BNLCC (P = 0.003). The optimum cutoff value of plasma sPLA2-IIa by ROC analysis was 2.4 ng/ml, which resulted in 64% sensitivity and 86% specificity for predicting lung cancer (AUC: 0.79; 95% CI: 0.69 ~ 0.90) (Table 3a). On the other hand, among 20 healthy donors, the levels of plasma sPLA2-IIa were undetecTable in 15 healthy donors, while those of the remaining 5 donors ranged up to 275 pg/ml (see Additional file 3). Age was not sig- nificantly associated with the level of plasma sPLA2-IIa in this healthy donor cohort (Table 1). ROC analysis of the plasma sPLA2-IIa levels in 96 lung cancer samples from the BNLCC and 20 healthy donor samples revealed an AUC as 1.0 (95% CI: 1.0 ~ 1.0) (Figure 2a). Further analysis revealed that plasma sPLA2-IIa was sig- nificantly higher in T2 stage lung cancer (n = 18) rela- tive to T1 stage lung cancer (n = 44) from the BNLCC (Two tailed t test: P = 0.005) (Figure 1b and see Addi- tional file 1). The optimum cutoff value of plasma sPLA2-IIa by ROC analysis was 2.4 ng/ml, which resulted in 67% sensitivity for predicting T2 stage lung cancer (AUC: 0.86; 95% CI: 0.65 ~ 0.96), as compared with 45% sensitivity for predicting T1 stage lung cancer (Table 3a and Figure 2c). High levels of plasma sPLA2-