How to collect animal specimen for experiments?

Collecting and storing animal tissue samples properly is crucial for accurate experimental results. Whether you're conducting histology, molecular analysis, or other research, following specific guidelines ensures sample integrity.

Tissue Sample Collection and Fixation

For most tissue samples, immediate fixation after removal from the body is essential to prevent degradation.

• Fixative Choice and Volume: The standard fixatives are 10% neutral formalin or 4% paraformaldehyde. Avoid using alcohol for fixation unless specifically required. The volume of fixative should be generous, typically 10 to 15 times the sample volume.
• Container Selection: Do not use V-bottom, slender, or small tubes for fixation, as these can hinder proper fixation.
• Large Specimens: For larger tissue specimens, cut them into smaller pieces before fixation to ensure the fixative penetrates thoroughly and prevents self-corruption in the center. A good general guideline for tissue block size is no more than 2.0cm x 2.0cm x 0.5cm.
• Special Tissues: Some tissues require specific fixatives to prevent alterations that could affect subsequent testing. Always consult your experimental protocols for these cases.
• Gastrointestinal Contents: When dealing with stomach and intestinal contents, do not squeeze the tissue, as this can damage the mucosa. Instead, carefully inject 10% neutral formalin or 4% paraformaldehyde into the stomach and cecum for initial fixation. After about half an hour, cut them open, wash with 10% sodium chloride solution, and then fix them again.
• Floating Tissues: For light tissues like skin or lungs that tend to float, you can gently place a wet cotton ball on top of the fixative to keep the tissue submerged. Alternatively, for whole lung tissue, inject the fixative into the main bronchus to flush and fill the entire lung before placing it in the fixative.
• Rinsing: Before immersing tissues in fixative, rinse them with saline to avoid insufficient fixation.

Frozen Section Specimens

If your experiment requires frozen sections, the tissue must be freshly collected.

• Processing: After removal, embed the tissue with OCT (Optimal Cutting Temperature compound) and immediately store it at -80℃.
• Transportation: During transport for inspection, ensure there's sufficient dry ice or ice packs to maintain the low temperature.

General Organ Collection Requirements

Adhering to these routine collection requirements will optimize your samples:

• Anesthetize and Bleed Animals: Animals should be anesthetized and bled before sampling to prevent excessive blood in the tissue, which can interfere with proper fixation.
• Prompt Fixation/Freezing: Once the animal dies and the tissue is removed, it must be fixed or frozen as soon as possible (within 10 minutes) to prevent tissue autolysis.
• Consistent Sampling: Collect samples from the same organ and the same part to ensure comparability of results in later experiments.
• Sharp Instruments: Use sharp instruments when sampling to avoid deforming or damaging the tissue through squeezing or scraping.

Storing Tissue Samples

• Fixed Samples: Tissue fixed with 10% neutral formalin or 4% paraformaldehyde can typically be stored at room temperature. Ensure the fixative solution completely covers the sample.
• Frozen Samples: Frozen section specimens embedded in OCT should be stored at -80℃.
• Refrigerated Samples: Samples stored at 4℃ (e.g., in a specific buffer or media) should be kept refrigerated. Avoid freezing the fixative itself during storage.

Mailing Animal Tissue Samples

Proper packaging and labeling are critical when mailing animal tissue samples to ensure their integrity and compliance with shipping regulations (e.g., UN3373 bag for biological substances, Category B).

Fixed Samples (Room Temperature Shipping): Tissue samples, tissue wax blocks, and non-fluorescent stained slides fixed with fixative can be sent at room temperature. When packaging slides, use paper towels or bubble wrap to prevent shock.

• Refrigerated Samples (4℃): Samples stored at 4℃ should be transported with ice packs. Do not allow the fixative to freeze during transport.
• Fluorescent Stained Slides: Place these slides in a slide box at low temperature and away from light. Include paper towels or bubble wrap for shock protection. Wrap the box with a layer of tin foil to avoid light exposure. Use sufficient ice or dry ice for low-temperature transport.
• Frozen Section Samples: These require ample dry ice and should be packaged in an insulated foam box to maintain their frozen state during transit.
• Documentation: All submitted specimens must include comprehensive information: species, specimen type, ex vivo time, fixation time, fixation type, and tissue location. This information must be clear and consistent with any accompanying paper documentation. Confirm that the tissue is completely fixed; if unsure, contact the pathology laboratory staff for guidance.
• Sample Size: Avoid submitting samples that are too small or thin (e.g., less than 1mm thick), as they are easily lost during processing.