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The Simplest and Quickest Method for T Cell Isolation from Mouse Spleen

April 2021

Spleen Cell Isolation

Just as a gold mine has different areas with higher concentrations of gold, different parts of the body have different levels of T cells. In a mouse, the spleen cell culture is composed of around 100 million splenocytes. Splenocytes are mononuclear white blood cells (WBCs) derived from or situated in the spleen. T cells typically comprise roughly 25% of the total splenocyte population.

While murine (relating to or affecting mice) T cells cannot be used to treat humans directly because of substantial biological differences, humans and mice have a largely similar genetic code. This allows scientists to use murine cells as a reference point for how human cells may react in various situations. Although not a perfect match, the accessibility and ease of using murine splenocyte cells make mice a common research subject for numerous areas of research, including immunology.

Splenocyte Breakdown

The spleen of a mouse contains a multitude of white blood cells that can be used for research, including:

  • CD4+ Helper T Cells (~16%) – Help the adaptive immune response by locating and identifying harmful pathogens
  • CD8+ Killer T Cells (~8%) – Release antibodies to eradicate pathogens
  • B Cells (~50%) – Present antigens and secrete cytokines to bolster immune response, can differentiate further into B cell subclasses
  • NK Cells (~1-2%) – Destroy cells that can’t be recognized by other lymphocytes
  • Granulocytes (~1-2%) – Help to destroy pathogens at infection site
  • Monocytes (~3-5%) – Move to infection site, turn into dendritic cells or macrophages
  • Dendritic Cells (~1-3%) – Signal immune system to begin response
  • Macrophages (~1-2%) – Responsible for disposing of cellular debris and waste

Methods of Murine T Cell Purification

T cell isolation from splenocytes can be done in a multitude of ways with a multitude of products. Depending on the target population, scope of cells, and desired next steps, the chosen isolation method can make a large difference in downstream results.

Magnetic Beads

One cell separation technique for T cell isolation from spleen cells is magnetic activated cell sorting, or MACS. A MACS protocol binds magnetic beads to target cells, then uses a magnetic field to separate the bound targets from the remainder of the sample.  This process is well-known and widely used but requires equipment and consumables (as well as trained personnel) to carry out. MACS is volume-limited and can only process small samples at a time, making it difficult to scale. It can also be harsh on delicate cells, and issues with non-specific binding and limited sensitivity can make it challenging to recover enough cells of low abundance when enriching for rare cell types.

Sorting with FACS

FACS, or fluorescence activated cell sorting, is another approach to cell separation. This method uses a machine called a flow cytometer to sort cells based on their physical properties. By labeling target cells with fluorescent markers, the machine can route them to different groups after scanning them with a laser and sensor.

FACS is extremely beneficial when the goal is to sort out multiple target populations. If the goal is to isolate two or more separate groups of cells, FACS is one of the most effective methods at performing this process. Results can be further optimized through effective sample preparation prior to sorting to remove unwanted contaminants. FACS is less accessible than other methods of cell separation, due to the time required to perform a sort and the expensive specialized equipment required. For routine sample processing, like murine T cell isolation, other approaches can be faster and easier.

Akadeum’s Microbubble Technology

Akadeum is revolutionizing the way separations are performed. Buoyancy-Activated Cell Sorting (BACS), Akadeum’s patented approach to separation, offers a novel way to isolate the target of interest using tiny floating particles we call microbubbles. Our technology combines the specificity of affinity molecules such as antibodies with the power of gravity in a workflow that’s fast, easy, ad effective while being exceptionally gentle on delicate targets, like immune cells of low abundance.

Our buoyant approach to separation eliminates many of the existing technical hurdles that are limiting improvements or effectiveness of processes today. Unlike processes like MACS or FACS, microbubbles do not have the same volume and equipment restrictions. Akadeum’s microbubble are gently mixed into the sample where they engage the targets before isolating those targets through floatation-based separation. It’s that simple! We’ve taken technology that traditionally requires additional equipment to accomplish this separation and put it into a single container where it’s self-separating. The fast and easy microbubble workflow takes place directly in the sample container and requires no additional equipment. Critically, this process is exceptionally gentle and maintains the health and physiology of cells of interest.

Isolation of Lymphocytes from Mouse Splenocyte Samples with BACS

Akadeum offers a portfolio of Mouse Immunology Kits that enrich for specific immune cells of interest from mouse splenocyte samples. The Akadeum microbubble enrichment protocol enables quick and easy sample preparation that takes about 30 minutes from start to finish without additional equipment or expensive consumables. The end result is a high-purity population of cells of interest that are ready for downstream use. In fact, using one of our microbubble kits prior to cell sorting can greatly reduce sort times, saving time and money while delivering a highly viable final population. Available microbubble kits for immune cell isolation from mouse splenocyte samples include:

  • Mouse B Cell Isolation Kit
  • Mouse T Cell Isolation Kit
  • Mouse CD4+ T Cell Isolation Kit
  • Mouse Naïve T Cell Isolation Kit
  • Mouse Naïve CD4+ T Cell Isolation Kit

If you’re interested in leveraging the power of microbubbles to overcome long-standing headaches in sample preparation in your lab, contact our team today. We’d love to hear more about your specific application and to find out whether microbubbles could be the solution you’ve been looking for.

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