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Tips and Tricks for Properly Handling 6-Well Cell Culture Plates

In the realm of cell biology and life sciences, cell cultivation is an indispensable technique that allows researchers to study and manipulate cells in a controlled environment. Among the various tools available, 6-well cell culture plates are a staple for many experiments. These plates are essential for the growth and maintenance of cells, but improper handling can lead to unreliable results. In this post, we will explore some key tips and tricks to help you master the art of properly handling these cellular cultivation plates.

1. Selecting the Right 6-Well Cell Culture Plate

Before you even start handling the cellular cultivation plate, it’s crucial to select the right one for your specific experiment. Consider the following factors such as:

  • Material: 6-well cell culture plates are typically made of either polystyrene or glass. Polystyrene plates are suitable for most experiments due to their transparency and compatibility with a wide range of cell types. Glass plates are used for specialized applications that require optical clarity, such as microscopy.
  • Coating: Some experiments may require specialized coatings, such as collagen, fibronectin, or poly-lysine, to facilitate cell adhesion and growth. Make sure to choose a plate with an appropriate coating if needed.
  • Lid Options: Consider whether you need a plate with a lid or a non-lidded version. Lidded plates help maintain sterility and prevent contamination, but non-lidded plates may be preferable for certain applications.

2. Aseptic Technique

Maintaining a sterile environment is paramount in cell cultivation work. Always work in a laminar flow hood, biosafety cabinet, or a clean, controlled laboratory space. Follow these guidelines for maintaining aseptic technique:

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  • Put on the proper personal protection gear, such as safety goggles, lab coats, and gloves.
  • Disinfect your work area and all materials with 70% ethanol or an appropriate disinfectant.
  • Flame-sterilize any tools, such as forceps or pipettes, that come into contact with the culture plates.

3. Handling and Seeding Cells

When handling 6-well cell culture plates during cell seeding, follow these best practices:

  • Gently pre-warm your culture medium to the appropriate temperature to prevent temperature shock to the cells.
  • Aspirate the old medium from the plate, being careful not to touch or disrupt the adherent cells.
  • When adding new cells or reagents, use a slow and controlled pipetting technique to avoid damaging the cells or creating air bubbles.
  • For even distribution of cells, use a zig-zag motion while pipetting the suspension into each well.
  • After seeding, gently swirl the plate to evenly disperse the cells, and then place it in an incubator under the appropriate conditions.

4. Proper Incubation

Proper incubation conditions are crucial for cellular cultivation success. Follow these tips:

  • Maintain the correct temperature, humidity, and CO2 levels in the incubator, as specified for your specific cell type.
  • Avoid frequently opening the incubator door to prevent temperature and CO2 fluctuations.
  • Regularly check the pH of the culture medium and replenish it with fresh medium as needed to maintain proper cell growth conditions.

5. Preventing Contamination

Contamination can ruin weeks of hard work. To prevent contamination in your 6-well cell culture plates:

  • Keep the cell culture area clean and free of clutter.
  • Always work with sterile materials and equipment.
  • Regularly check for contamination under a microscope, which may include bacterial or fungal growth.
  • Handle the culture plate lids carefully, and only open them when necessary.
  • Minimize the time the culture plates are outside of the incubator to reduce the risk of airborne contaminants settling in.

6. Feeding and Maintaining Cells

Maintaining the health of your cultured cells is an ongoing process. Here are some tips for feeding and maintaining them:

  • To add fresh vitamins and minerals and get rid of waste materials, switch up the culture medium frequently.
  • Use aseptic techniques when changing the medium, and be mindful of the volume you add to each well to prevent overfeeding or underfeeding the cells.
  • Keep a careful record of your feeding schedule and any observations about the cell culture’s health.
  • If you observe cell detachment or irregular growth, troubleshoot the issue promptly.

7. Handling and Passaging Cells

Passaging is the process of subculturing cells to prevent over-confluency and maintain their health. When passaging cells in 6-well plates:

  • Aspirate the old medium gently.
  • Wash the cells with an appropriate buffer or solution, depending on your experimental needs.
  • Use trypsin or another suitable enzyme to detach the cells from the culture plate.
  • Pay attention to the cell density when replating, as overcrowding can affect their growth.

8. Minimizing Evaporation

Cell culture plates can lose moisture through evaporation, which can affect the concentration of nutrients in the medium. To minimize evaporation:

  • Use humidified incubators to maintain proper humidity levels.
  • Ensure the culture plate lids fit snugly and are not damaged.
  • Consider the use of paraffin oil or plate sealers to further reduce evaporation.

9. Monitoring Cell Growth

Regular monitoring of cell growth is essential for successful cellular cultivation experiments. Use a microscope to check for the following:

  • Cell confluency: Determine when the cells have reached the desired density for your experiment.
  • Cell morphology: Examine the shape and appearance of the cells to identify any abnormalities.
  • Contamination: Regularly check for signs of contamination, such as bacterial or fungal growth.

10. Record Keeping

Maintain detailed and organized records of your cell culture work. This includes:

  • Cell source information.
  • Culture medium and supplements are used.
  • Passage number and dates of subculture.
  • Any issues or observations during the experiment.
  • Experimental results and outcomes.

Having well-organized records can help troubleshoot problems and ensure the reproducibility of your experiments.


Properly handling 6-well cell culture plates is a fundamental aspect of cellular cultivation work that can significantly impact the success of your experiments. Maintaining a sterile environment, following best practices for handling and storage, and paying attention to detail during each step of your experiments are key to obtaining reliable and reproducible results. By implementing the tips and tricks outlined in this article, you can master the art of cell cultivation and elevate the quality of your research. Remember, the devil is in the details, and excellence in cell culture is built upon a foundation of meticulous care and precision.

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