Biomedical journal · 2025
Elastase reduces background autofluorescence in ALK fluorescence in situ hybridization assays for lung cancers.
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Hsu Sheng-Chi, Hung Tsai-Hsien, Wu Hsiao-Chun, Ng Kwai-Fong, Chen Tse-Ching
Аннотация
BACKGROUND: Anaplastic lymphoma kinase (ALK) inhibitors have been effective in treating non-small cell lung cancers (NSCLC) with ALK translocation. However, high background autofluorescence in lung tissues interferes with fluorescence in situ hybridization (FISH) assays, masking molecular probe signals and hindering data interpretation.
MATERIALS AND METHODS: To reduce autofluorescence, NSCLC tissue sections were treated with various proteases, including collagenase types I, II, IV, and elastase, to determine the most effective enzyme. We then conducted ALK break-apart FISH assays on 120 NSCLC samples, comparing standard and novel pretreatment protocols.
RESULTS: Elastase was identified as the most effective enzyme for reducing autofluorescence while preserving nuclear integrity. The elastase-based pretreatment enabled clear FISH signal detection in all cases, reducing the retest rate from 86.7% to 0%. Furthermore, two additional ALK translocated cases were detected with elastase pretreatment, which were indeterminable with pepsin treatment alone.
CONCLUSIONS: This novel elastase pretreatment protocol addresses autofluorescence interference in lung tissues and can significantly improve the reliability of FISH assays for targeted therapy decisions.
MATERIALS AND METHODS: To reduce autofluorescence, NSCLC tissue sections were treated with various proteases, including collagenase types I, II, IV, and elastase, to determine the most effective enzyme. We then conducted ALK break-apart FISH assays on 120 NSCLC samples, comparing standard and novel pretreatment protocols.
RESULTS: Elastase was identified as the most effective enzyme for reducing autofluorescence while preserving nuclear integrity. The elastase-based pretreatment enabled clear FISH signal detection in all cases, reducing the retest rate from 86.7% to 0%. Furthermore, two additional ALK translocated cases were detected with elastase pretreatment, which were indeterminable with pepsin treatment alone.
CONCLUSIONS: This novel elastase pretreatment protocol addresses autofluorescence interference in lung tissues and can significantly improve the reliability of FISH assays for targeted therapy decisions.