The most effective method for monitoring country-level drug resistance frequency and to implement the necessary control measures is the establishment of a laboratory-based surveillance system. The aim of this study was to summarize the follow up trend of the drug-resistant tuberculosis (TB) cases, determine the load of resistance and evaluate the capacities of laboratories depending on laboratory quality assurance system for the installation work of National Tuberculosis Laboratory Surveillance Network (TuLSA) which has started in Ankara in 2011. TuLSA studies was carried out under the coordination of National Tuberculosis Reference Laboratory (NRL) with the participation of TB laboratories and dispensaries. Specimens of TB patients, reported from health institutions, were followed in TB laboratories, and the epidemiological information was collected from the dispensaries. One isolate per patient with the drug susceptibility test (DST) results were sent to NRL from TB laboratories and in NRL the isolates were rechecked with the genotypical (MTBDRplus, Hain Lifescience, Germany) and phenotypical (MGIT 960, BD, USA) DST methods. Molecular epidemiological analysis were also performed by spoligotyping and MIRU/VNTR. Second-line DST was applied to the isolates resistant to rifampin. A total of 1276 patients were reported between January 1st to December 31th 2011, and 335 cases were defined as "pulmonary TB from Ankara province". The mean age of those patients was 43.4 ± 20 years, and 67.5% were male. Three hundred seventeen (94.6%) patients were identified as new cases. The average sample number obtained from pulmonary TB cases was 3.26 ± 2.88, and 229 (68.3%) of them was culture positive. DST was applied to all culture positive isolates; 90.4% (207/229) of cases were susceptible to the five drugs tested (ethambutol, isoniazid, pyrazinamide, rifampicin, streptomycin). Eight (3.5%) of the isolates were multidrug-resistant (MDR-TB), while no extensively drug-resistant strains were detected. MDR-TB is likely to occur in 63.3 times more among previously treated cases, and 73.3 times more in legal aliens. The achievement of therapy among pulmonary TB cases was 91.9%. Spoligotyping performed for 221 M.tuberculosis complex isolates, showed that all strains were clustered in nine groups. SIT 41 (105/221; 47.5%) was the most frequent spoligotype detected, and clustering rate based on MIRU-VNTR results were found as 16.3%. All of the clustered strains were sensitive while all of MDR-TB isolates showed specific MIRU-VNTR profiles. In conclusion, TuLSA studies started in Ankara in 2011 and the system is still expanding in the country. Our data obtained with TuLSA have been published as a regional surveillance data in the WHO Global Tuberculosis Report 2011, and as a national surveillance data in Global Tuberculosis Report 2012.