GSM9291475: A07; Homo sapiens; OTHER
Colorectal cancer (CRC) tissue specimens were collected from a patient with stage IV disease during surgical resection at Siriraj Hospital, Mahidol University, Thailand. Three preservation methods were used: formalin-fixed paraffin-embedded (FFPE), fresh frozen (FF), and snap frozen (SF). For FF tissues, specimens were cut into 2–3 mm pieces within 30 minutes of resection and immersed in chilled RNAlater solution. SF tissues were rapidly frozen in liquid nitrogen. FFPE tissues were fixed in formalin and embedded in paraffin according to standard clinical practice. Sections (5 µm) from all tissue types were mounted on Superfrost Plus slides for spatial transcriptomic analysis. RNA integrity was verified (DV200 > 50%) before library preparation. Barcoded oligonucleotides released from defined tissue areas were collected and sequenced on an Illumina NovaSeq 6000 following NanoString GeoMx DSP Human Whole Transcriptome Atlas (WTA) workflow guidelines. Spatial whole-transcriptome libraries were constructed using the NanoString GeoMx Digital Spatial Profiler (DSP) Human Whole Transcriptome Atlas (WTA) workflow. Tissue sections were hybridized with UV-photocleavable oligonucleotide probes complementary to target mRNA transcripts. Following region-of-interest (ROI) selection, oligonucleotide tags were released from each compartment (PanCK⁺, CD45⁺, and PanCK⁻/CD45⁻) by UV illumination and collected into separate wells. The released barcodes were subjected to PCR amplification and indexing using the GeoMx WTA Library Preparation Kit, following the manufacturer's protocol. Final libraries were purified, quantified, and quality-checked before sequencing on an Illumina NovaSeq 6000 platform.