Steroid sulfatase (STS) is a membrane-bound microsomal enzyme that hydrolyzes various alkyl and aryl steroid sulfates, leading to the in situ formation of biologically active hormones. The entire human STS gene spans over approximately 200kbp of which the first 100kbp include the regulatory region, while the STS-coding region is located downstream. Previous studies indicated that STS expression, in different human tissues, could be regulated by at least six different promoters associated with alternative first exons. Here, we describe two new splicing patterns: the first, found in the prostatic cell line PC3, is based upon a partially coding new first exon (0d) that is spliced to a new second exon (1e). The second variant was found in the ovary and it is characterized by the novel splicing of the untranslated exon 0b to exon 0c, which is then spliced to the common exon 1b. We also report the results of a multiplex ligation-dependent probe amplification (RT-MLPA) analysis for the simultaneous detection, in qualitative and/or semi-quantitative terms, of the transcription patterns of STS in different tissues.