How to count cells with the Neubauer Counting Chamber

Summary

The Bower Counting Chamber offers a quick and effective method for manually counting cells in a solution, despite the rise of automated devices. The chamber features nine squares, each with a volume of 0.1 microliter, and cells are counted within these squares under a microscope. The process includes cell dilution and sample application, followed by counting cells while adhering to specific rules about counting cells on boundaries. Finally, a calculation converts the count into cells per milliliter, with an example illustrating a final count of 1 million cells per milliliter. This technique is still widely used for cell counting in laboratory settings.

Takeaways

• 🧫 The Neubauer counting chamber enables quick and simple manual determination of cell numbers in a solution.
• ⚙️ Despite modern automated cell counters, manual counting with a Neubauer chamber remains widely used in laboratories.
• 📏 Each sample spot on the Neubauer slide measures 3×3 mm and contains nine squares, with each large square further divided into 16 smaller boxes.
• 🔬 One of the nine large squares used for counting has an area of 1 mm², and the chamber’s depth is 0.1 mm, giving each square a volume of 0.1 µL.
• 🧍‍♀️ To count cells, the cells are first detached from the culture flask and can be diluted for easier counting if needed.
• 💧 A common dilution example is mixing 1 mL of cell suspension with 9 mL of medium or water, giving a 1:10 dilution (dilution factor = 10).
• 📸 About 10 µL of the diluted sample is then applied to the counting chamber for microscopic observation.
• 📊 Counting follows a specific boundary rule: include cells touching the lower and right borders, exclude those touching the upper and left borders.
• 🔢 The average number of cells per counting square is calculated, and cell concentration per mL is determined using the formula: (average cells per square) × 10⁴ × dilution factor.
• 🧮 In the example given, 10 cells per square with a dilution factor of 10 result in a total of 1,000,000 cells per milliliter.
• 🎥 The video ends with a lighthearted note encouraging viewers to subscribe to reach one million subscribers, paralleling the cell count example.

Q & A

What is the Bower counting chamber used for?

The Bower counting chamber is used to determine the number of cells in a solution, offering a traditional method of cell counting, even though modern automated devices are also available.

How is the Bower counting chamber designed for cell counting?

The Bower counting chamber features two spots for applying the sample, each measuring 3x3 mm. These spots consist of nine large squares, each 1x1 mm, divided into 16 smaller boxes for counting cells.

What is the volume of one of the large squares in the Bower counting chamber?

One of the large squares in the Bower counting chamber has a surface area of 1 square millimeter and a volume of 0.1 microliter, due to the 0.1 mm distance between the slide and the coverslip.

Why is the height of the solution important when using the Bower counting chamber?

The height of the solution is important because it directly affects the volume of the sample being counted. In the Bower counting chamber, the height is 0.1 mm, determining the volume of 0.1 microliter for each of the large squares.

What is the purpose of diluting the cell suspension in the process?

Diluting the cell suspension, such as in a 1:10 ratio (1 ml of cells in 9 ml of water or medium), is optional but helps especially when there is a high concentration of cells, making counting more manageable.

How do you count cells in the Bower counting chamber?

To count cells, you place the sample in the counting chamber and observe it under a microscope. The cells in the highlighted squares are counted. The rule is to count cells touching the lower or right boundaries but exclude those touching the upper or left boundaries.

What is the counting rule for cells that touch the boundaries of the squares?

Cells touching the lower or right boundary of a square are counted, while cells touching the upper or left boundary are not counted.

How do you calculate the number of cells per milliliter from the Bower counting chamber?

First, calculate the average number of cells per square. Then, multiply that number by 10,000 (to convert to cells per milliliter, as 1 ml = 10,000 times 0.1 microliter). Finally, multiply by the dilution factor (e.g., 10 in the case of a 1:10 dilution).

Why is the dilution factor necessary in the calculation?

The dilution factor is necessary because it accounts for any dilution of the original cell suspension. In the example of a 1:10 dilution, the factor of 10 ensures the final count reflects the concentration in the undiluted sample.

What was the final cell concentration in the example given in the video?

In the example, the final cell concentration was 1 million cells per milliliter, based on a count of 10 cells per square, multiplied by 10,000 (for volume conversion) and the dilution factor of 10.