Leishmania infantum is the most common cause of visceral leishmaniasis (VL) in Iran, where mainly the patients are children under the age of 5 years. Timely, less invasive, and accurate diagnosis and proper treatment of the disease are necessary. This retrospective study aimed to search for a less invasive but robust algorithm on VL diagnostic tests in children. Four hundred and fifteen patients with clinical suspicion of VL, 50 healthy children from VL endemic areas, 46 healthy individuals from non-endemic VL areas, and 47 non-VL diseases were tested using three diagnostic tests: indirect immunofluorescent antibody test (IFAT), rK39-rapid diagnostic test (rK39-RDT), and quantitative PCR (qPCR). One hundred and two suspected VL cases were positive in at least one test and were cured after receiving appropriate treatment. Of these 102 VL patients, 94 were positive in qPCR, 84 in IFAT, and 79 in rK39-RDT. None of the tests detected all the patients, but overall, qPCR is capable of detecting more VL patients than serological tests, i.e., 92.2%, compared to IFAT, 82.4%, and rK39, 77.5%. There was only a significant difference between the sensitivity of qPCR and rK39-RDT (p = 0.024). The specificity was 100% for qPCR and IFAT (≥128) and 98.6% for rK39-RDT. qPCR alone is capable of detecting most of the VL-suspected children. Serological tests like IFAT and rk39-RDT are recommended to increase the overall sensitivity of detection in patients with a negative molecular test. Combining qPCR with a serological test (IFAT or rK39-RDT) can help diagnose 98% of VL. In laboratories without molecular facilities, we recommend testing with the combination of rK39-RDT and IFAT yielding a combined sensitivity of 93.1% equivalent to that of qPCR in our study.