Yes, you can kill bacteria without lysing them. Heat fixation is a common method and works well for basic microscopy, though it might slightly alter cell shape. If you want an alternative, chloroform vapor is a good option. Just place a chloroform-soaked paper in a sealed container, set your sample above it (not touching the liquid), and let it sit for about 20 minutes. This will kill the bacteria while keeping their structure intact. For more precise shape preservation, chemical fixatives like formaldehyde can also be used, but they require careful handling.

Let me know what kind of microscope you're using, I can maybe help narrow it down further.

Need to know what bacterium

Beyond heat-fixation, chemical fixation is the best method to kill bacteria while preserving their morphology for microscopy in a clean room. Chemical fixatives cross-link proteins, stabilizing cellular structures with less distortion than heat.

The choice of chemical fixative depends on the type of microscope and staining you intend to use.

Aldehyde fixation

Aldehydes are excellent cross-linking fixatives that preserve fine cellular details.

Paraformaldehyde (PFA)

• Mechanism: A gentle fixative that cross-links proteins, maintaining cellular structure and integrity with minimal shrinkage. It is widely used for fluorescence microscopy because it preserves antigenicity for immunostaining.
• Protocol (example):Prepare a fresh 4% PFA solution in a suitable buffer like phosphate-buffered saline (PBS) with a pH of ~7.4. Heating is required to dissolve the paraformaldehyde powder. Carefully create a bacterial smear on a clean microscope slide. Immerse the slide in the PFA solution for 10–20 minutes at room temperature. Wash the slide thoroughly with PBS to remove residual PFA before taking it to the clean room.

Glutaraldehyde (GA)

• Mechanism: A stronger and faster-acting cross-linking agent than PFA. It is the gold standard for preserving fine ultrastructure in electron microscopy (EM), but its strong autofluorescence makes it unsuitable for fluorescence microscopy.
• Protocol: A typical concentration is 2.5% GA in a buffer like 0.1 M cacodylate. For your application, you can fix the bacterial smear on a slide for 30–60 minutes, then wash it thoroughly with buffer.

Organic solvent fixation

Organic solvents like methanol, ethanol, and acetone coagulate and precipitate proteins. They are fast-acting but can remove lipids and cause more cellular shrinkage than aldehydes.

Cold methanol

• Mechanism: Removes lipids and dehydrates the cells, killing them by denaturing and precipitating proteins. The cold temperature minimizes the risk of altering morphology.
• Protocol (example):Create the bacterial smear on a slide. Rinse briefly with PBS to remove excess media. Incubate the slide in 100% ice-cold methanol for 5–15 minutes at -20°C. Wash the slide gently three times with PBS before observation.

Chloroform vapor fixation

This method kills bacteria without causing significant cell wall damage, making it suitable for preserving basic morphology.

Chloroform vapor protocol