How To Prepare A Wet Mount Slide

How to Prepare a Wet Mount Slide: A Comprehensive Guide

Creating a wet mount slide is a fundamental technique in microscopy, allowing you to observe living organisms and specimens in their natural aqueous environment. This detailed guide will walk you through the process, covering various aspects from choosing the right materials to troubleshooting common issues. Whether you're a student, researcher, or simply curious about the microscopic world, mastering this technique will unlock a whole new level of exploration.

Understanding the Wet Mount Technique

A wet mount slide involves placing a specimen in a drop of liquid on a microscope slide, then covering it with a coverslip. The liquid acts as a mounting medium, keeping the specimen hydrated and preventing it from drying out during observation. This simple yet powerful technique allows for observation of living microorganisms, cells, and other delicate specimens without the need for complex preparation or staining procedures.

Advantages of Wet Mounts:

• Observation of living specimens: Unlike stained slides, wet mounts allow you to observe the natural movement and behavior of living organisms.
• Simplicity and speed: Preparation is quick and easy, requiring minimal equipment and expertise.
• Cost-effectiveness: The materials are readily available and inexpensive.
• Versatility: Suitable for observing a wide range of specimens, from pond water samples to prepared biological specimens.

Disadvantages of Wet Mounts:

• Limited longevity: The specimen can dry out, especially with prolonged observation, affecting the quality of the image and potentially damaging the specimen.
• Potential for artifacts: Air bubbles or debris can interfere with observation.
• Movement of the specimen: The specimen can move around, making observation challenging. This can be mitigated through techniques like using a sealant or reducing the amount of liquid.
• Difficult to permanently store: Wet mounts are generally not suitable for long-term storage.

Essential Materials and Equipment

Before you begin, ensure you have the following materials and equipment:

• Microscope slides: Standard microscope slides made of glass are essential. Clean slides are crucial for optimal viewing.
• Coverslips: These thin, square pieces of glass are used to cover the specimen and prevent it from drying out. Sizes vary; #1 coverslips are commonly used.
• Pipettes or Droppers: For accurately transferring the mounting medium and the specimen onto the slide. Pasteur pipettes are ideal for smaller specimens.
• Tweezers: For carefully handling specimens, especially small ones or those that require delicate manipulation.
• Mounting medium: The choice of mounting medium depends on the specimen. Distilled water is a common choice for many biological specimens. Other options include saline solution, glycerin, or specialized mounting media for specific applications.
• Specimen: This is the object of your observation. This could be anything from a drop of pond water teeming with microorganisms to a prepared sample of cheek cells.
• Lens paper: Used for cleaning microscope slides and coverslips.
• Microscope: Naturally, you'll need a microscope to view your wet mount slide.

Step-by-Step Guide to Preparing a Wet Mount Slide

Follow these steps for optimal results:

1. Preparing the Slide:

• Clean the slide: Ensure your microscope slide is clean and free from dust or fingerprints. Use lens paper to gently wipe the slide.
• Add the mounting medium: Place a small drop (approximately 1-2 mm in diameter) of the mounting medium – usually distilled water – in the center of the slide. Too much liquid will cause the coverslip to float, making observation difficult. Too little may cause the specimen to dry out quickly.

2. Adding the Specimen:

• Transfer the specimen: Using tweezers or a pipette, carefully transfer your specimen to the drop of mounting medium. Avoid introducing air bubbles. If working with a liquid sample like pond water, simply use a pipette to transfer a small amount. For solid specimens, carefully position them within the drop.

3. Applying the Coverslip:

• Lower the coverslip: Gently lower the coverslip onto the specimen at a 45-degree angle. This minimizes the formation of air bubbles. Slowly lower the coverslip to avoid trapping air bubbles. If air bubbles are present, gently tap the coverslip to try to dislodge them.
• Avoid trapping air: Use a slow and steady approach when placing the coverslip. If air bubbles are present, attempt to gently maneuver them to the side of the coverslip.

4. Observing under the Microscope:

• Place the slide: Carefully place the wet mount slide on the microscope stage.
• Focus the microscope: Begin with the lowest magnification objective lens and slowly adjust the focus until the specimen is clearly visible. Gradually increase the magnification as needed.

Choosing the Right Mounting Medium

The choice of mounting medium is crucial for the success of your wet mount. Here's a guide to common options:

• Distilled Water: The simplest and most commonly used mounting medium. Ideal for observing living organisms in their natural environment. However, it evaporates quickly, limiting observation time.
• Saline Solution: A solution of salt and water, which is often used for observing specimens that are sensitive to changes in osmotic pressure. Similar evaporation concerns to distilled water.
• Glycerin: A viscous liquid that slows down evaporation, allowing for longer observation periods. It can, however, affect the refractive index, potentially altering the appearance of some specimens.
• Specialized Mounting Media: For specific applications or organisms, commercial mounting media with added preservatives or buffers are available.

Troubleshooting Common Wet Mount Problems

Despite the simplicity of the technique, several challenges can arise. Here are common issues and their solutions:

• Air Bubbles: These are a common problem, particularly when adding the coverslip. Slow and careful application is key. Gently tapping the coverslip may dislodge them.
• Specimen Drying Out: This is more prevalent with water-based mounting media. Using a glycerin-based medium or sealing the edges of the coverslip with petroleum jelly can extend the observation time.
• Coverslip Movement: The coverslip may move around, especially with larger specimens. Adding a small amount of mounting medium can help to stabilize it.
• Specimen Too Thick: Thick specimens can make focusing difficult. Consider using a thinner specimen or sectioning the specimen to reduce its thickness.
• Low Contrast: If the specimen is difficult to see, try adjusting the microscope's lighting or staining the specimen, although staining is not compatible with observing living cells.

Advanced Wet Mount Techniques

For more advanced applications, you can consider these techniques:

• Ringing: Sealing the edges of the coverslip with a sealant (e.g., clear nail polish) helps to prevent evaporation and keeps the specimen from drying out. This is particularly useful for longer observation periods or for creating slides suitable for short-term storage.
• Using a Depression Slide: These slides have a recessed well, which helps to contain the specimen and mounting medium, making the preparation easier. Depression slides minimize the likelihood of a coverslip shifting.
• Using a Hanging Drop Slide: Instead of placing the specimen on top, a hanging drop technique suspends the sample from an inverted coverslip. This helps in observing the motility of microorganisms without disturbing their movement.

Conclusion

Preparing a wet mount slide is a straightforward yet essential technique in microscopy. By carefully following the steps outlined in this comprehensive guide, and understanding the crucial details of selecting the appropriate mounting medium and addressing common problems, you can effectively observe a wide variety of specimens. Mastering this technique will undoubtedly enhance your microscopic exploration and open up new avenues of discovery in the fascinating world of microscopy. Remember that practice makes perfect! The more wet mounts you create, the more confident and proficient you will become in this valuable laboratory skill.