Klinefelter syndrome (KS) is the most common chromosomal syndrome, causing infertility in men and leading to non-obstructive azoospermia. Previous studies on mosaicism have shown contradictory results on its correlation with both serum hormone levels and the presence of spermatozoa in the ejaculate of KS, KS-like, and non-KS-like infertile patients. So, the present study was designed to detect low-grade mosaicism in the peripheral blood lymphocytes and buccal mucosa cells of 14 KS and 8 KS-like patients by using fluorescence in situ hybridization (FISH) and to investigate its correlation with luteinizing hormone (LH), follicle-stimulating hormone (FSH), and testosterone (T) levels, testicular volume, and semen analysis compared with 10 normal healthy fertile men. Our results indicated that mosaicism was only found in 42.9 % of the KS patients and completely absent in all KS-like patients. Moreover, mosaicism has led to complete azoospermia and non-significant differences in both hormone levels and testicular volume between mosaic and non-mosaic KS patients. All KS patients demonstrated significant differences in both hormone levels and testicular volume compared with normal men. Conversely, they revealed non-significant differences in hormone levels and significant differences in testicular volume compared with KS-like patients. Additionally, the KS-like patients exhibited non-significant variations in both LH and FSH levels and significant variations in T level and testicular volume compared with normal men. Moreover, all KS-like patients had azoospermia, except for one patient who showed oligozoospermia. Therefore, no correlations were found either between mosaicism and serum hormone levels or with testicular volume and semen analysis.