An alternative transcript variant of SLC39A14, caused by pre-mRNA splicing, was recently suggested as a biomarker for colorectal cancer (CRC). In our study, we have validated the cancer-specific splicing pattern of the mutually exclusive exons 4A and 4B in altogether 244 colorectal tissue samples. Exon-specific quantitative RT-PCR analyses across 136 Stage I-IV CRC samples and 44 normal colonic mucosa samples showed complete cancer-specificity, as well as 94% sensitivity of SLC39A14-exon4B relative to SLC39A14-exon4A expression. However, across 20 samples from a range of healthy tissues, 18 expressed the CRC variant. This was true also for ten benign lymph nodes, demonstrating that the cancer-specificity is mainly confined to the colon and rectum. Hence, clinical use of SLC39A14-exon4B as a detection marker for CRC other than in samples taken from the bowel wall is diminished. Prognostic value by detection of metastasis to lymph nodes is also abated, elucidating an important pitfall to biomarker discovery. However, analyses of ten nondysplastic biopsies from patients with active inflammatory bowel disease showed negative results in seven samples and only weakly positive results in three samples, suggesting value of SLC39A14-exon4B as a marker to distinguish CRC from other pathologic conditions of the colon. In conclusion, the SLC39A14-exon4B transcript variant is a CRC biomarker with high sensitivity and organ-confined specificity. Further use of the transcript and its encoded protein isoform should be explored in an organ-confined context.Colorectal cancer (CRC) is one of the most prevalent and lethal cancers worldwide. 1 The only biomarker approved for routine clinical use in CRC is serum levels of carcinoembryonic antigen to aid in postoperative monitoring of Stage II and III disease, as well as metastatic disease during systemic therapy. 2 Recently, solute carrier family 39 (zinc transporter), member 14 (SLC39A14) was proposed as a potential biomarker for CRC, with indications of applicability also in gastric and lung cancers. 3 Exons 4A and 4B of this gene are mutually exclusive and cancer-specific alternative splicing results in an increased exon 4B/4A ratio in both carcinomas and adenomas. Colorectal, gastric and lung cancers are known to have an aberrantly activated WNT-signaling pathway. [4][5][6] Hence, SLC39A14-exon4B was proposed as a detection biomarker in cancer types found to have an activated WNT-signaling pathway. 3 For a biomarker to be considered for clinical use, its applicability should be carefully validated in independent datasets by independent researchers. 7 No such validation studies have been reported for SLC39A14.In our study, we have confirmed cancer-specific alternative splicing of SLC39A14, i.e., high relative expression of SLC39A14-exon4B compared to SLC39A14-exon4A, in CRC by exon microarray and quantitative reverse transcription PCR (qRT-PCR) analyses. However, we found this specificity to be organ-confined, as the biomarker was positive also for healthy tissues from other organ...