What are the recommended OCT embedding procedures and precautions?

Answer:
SeekSpace supports two primary OCT embedding workflows.

Method 1: Liquid nitrogen + isopentane embedding (recommended)

1. Pre-cool OCT compound and related materials on wet ice for at least 30 minutes.
2. Remove surface moisture from tissue and complete embedding as quickly as possible (typically within 30 minutes).
3. Add a small amount of OCT to the mold base, place the tissue with the region of interest parallel to the mold bottom, then cover with OCT while avoiding bubbles.
4. Rapidly freeze the mold in isopentane. Once the OCT turns completely white, transfer the sample to a sealed −80°C freezer.

Method 2: Dry-ice-assisted embedding (optional)

1. Pre-cool OCT compound and tools on wet ice or dry ice for at least 30 minutes.
2. Rapidly freeze tissue on dry ice until the tissue surface turns white.
3. Add a small amount of OCT to the mold base, place the frozen tissue inside, cover with OCT, maintain the region of interest parallel to the mold base, then freeze until the OCT turns white before storage at −80°C.

Important precautions

• Tissue cross-sectional area should remain between 3 mm × 3 mm and 5.5 mm × 15 mm.
• The region of interest thickness should exceed 200 μm.
• Label sample orientation and dimensions before embedding.
• Remove excess tissue surface moisture to reduce ice crystal formation.
• During liquid nitrogen + isopentane embedding, do not fully immerse the mold in isopentane, as this may wrinkle the OCT surface.
• During dry ice embedding, ensure uniform cooling and keep regions of interest from multiple tissues on the same plane.

Core principles

• Maintain rapid low-temperature processing to minimize RNA degradation.
• Preserve a flat region of interest to support efficient barcode and RNA capture.
• Avoid bubbles and ice crystals to maintain tissue integrity.