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Ultra-rapid microwave-stimulated tissue processing with a modified protocol incorporating microwave fixation
Zenobia A M Hafajee1, Anthony S-Y Leong
1Immunohistology Unit, Hunter Area Pathology Service, and Discipline of Anatomical Pathology, University of Newcastle, Newcastle NSW 2310, Australia.
Aims:
To develop an ultra-rapid microwave (MW)-stimulated histoprocessing protocol that incorporates MW fixation and produces consistent, high quality sections.
Methods:
A range of fresh autopsy tissues was divided into three groups composed of equal numbers of small and large tissue blocks. Group 1 tissues were fixed for 8 hours in 4% buffered formaldehyde and processed in a conventional tissue processor through a 15-hour cycle. Group 2 tissues were processed in a MW histoprocessor according to the manufacturer's recommended protocol of irradiation in a proprietary reagent before embedding in paraffin. Group 3 tissues were processed with our protocol that incorporated the addition of MW fixation in 4% buffered formaldehyde and MW irradiation in isopropyl alcohol after irradiation in the proprietary reagent.
Results:
The manufacturer's protocol resulted in 'grey' and 'wet look' artefacts in large tissue blocks. These artefacts did not occur in our protocol which produced sections that were indistinguishable from those obtained with conventional 23-hour processing. Histochemical and immunohistochemical stains were also indistinguishable and the tissue blocks cut very smoothly.
Conclusions:
The incorporation of an additional step to ensure adequate tissue fixation and one to ensure optimal dehydration and clearing resulted in a MW histoprocessing protocol that produced consistent results for both large and small tissue blocks. Paraffin-impregnated blocks can be produced very rapidly from fresh small and large tissue blocks in 45 and 100 minutes, respectively.
Insights
This study introduces an ultra-rapid microwave (MW) histoprocessing protocol for consistent, high-quality tissue sections. The new method significantly reduces processing time for both small and large tissue blocks.
Area of Science:
- Histopathology
- Biotechnology
- Medical Laboratory Science
Background:
- Conventional tissue processing is time-consuming, often exceeding 24 hours.
- Existing microwave-assisted methods may introduce artifacts, especially in larger tissue samples.
- Optimizing histoprocessing is crucial for timely diagnostic and research applications.
Purpose of the Study:
- To develop an ultra-rapid microwave (MW)-stimulated histoprocessing protocol.
- To incorporate MW fixation for enhanced speed and quality.
- To achieve consistent, high-quality tissue sections from both small and large blocks.
Main Methods:
- Tissues were divided into three groups: conventional processing, manufacturer's MW protocol, and a novel MW protocol.
- The novel protocol included MW fixation in formaldehyde and MW irradiation in isopropyl alcohol.
- Comparison of processing times, artifact occurrence, and section quality.
Main Results:
- The manufacturer's MW protocol produced artifacts in large tissue blocks.
- The novel MW protocol yielded sections indistinguishable from conventional methods, without artifacts.
- Histochemical and immunohistochemical staining results were comparable to conventional processing.
Conclusions:
- The developed MW histoprocessing protocol ensures consistent, high-quality results for all tissue block sizes.
- This protocol significantly reduces processing time, enabling paraffin-impregnated blocks in as little as 45 minutes (small) and 100 minutes (large).
- The optimized protocol enhances efficiency in histopathological workflows.
