Preservation of Cell Surface Epitopes with STEMprep™ Mouse Tumor Dissociation Kit
- Document # TB136
- Version 1.0.0
- Jun 2025
Tumor tissues are inherently complex, made up of heterogeneous cell populations that express distinct cellular markers. Preserving these epitopes during tissue dissociation is critical for downstream applications such as flow cytometry, cell sorting, and cell separation—where marker integrity directly impacts data accuracy and cell characterization.
The STEMprep™ Mouse Tumor Dissociation Kit is specifically developed to address the challenge of maintaining epitope integrity during dissociation. Compatible with a wide range of tumor types, the kit enables the generation of high-yield, high-viability single-cell suspensions while minimizing enzymatic impact on surface epitopes. To assess this, we evaluated the sensitivity of over 200 surface markers to the STEMprep™ Mouse Tumor Dissociation Cocktail. The results confirm that the majority of epitopes remain preserved and detectable following treatment, enabling consistent characterization of tumor cells.
Method
Mouse spleen, lung, and bone marrow tissues were manually processed without enzymes to generate a single-cell suspension containing both immune and non-immune populations. These cells were blended with three tumor cell lines (4T1, B16, and CT26) to incorporate tumor-associated epitopes (see Figure 1). The single-cell suspension, comprising a diverse mix of cell types, was divided into two portions: one part was left untreated and labeled with CellTrace™ Violet (CTV) dye, and the other was treated with the STEMprep™ Mouse Tumor Dissociation Cocktail and incubated in the STEMprep™ Tissue Dissociator for 40 minutes at 37°C. The samples were then combined in equal parts and stained with the LEGENDScreen™ Mouse PE Kit and additional antibodies, along with an anti-mouse CD45 antibody, and analyzed by flow cytometry. The CTV dye allows distinction between enzyme-treated and untreated cells, while the CD45 antibody differentiates immune cells from non-immune cells or tumor cells. The effect of incubation with STEMprep™ Mouse Tumor Dissociation Cocktail on cell surface epitope integrity was evaluated by changes in fluorescence intensity and the percent positive population measured by flow cytometry.
Figure 1. Workflow to Assess Epitope Sensitivity to STEMprep™ Mouse Tumor Dissociation Cocktail.
Results
Assessed epitopes were categorized into three groups—stable, moderate, and sensitive—based on changes in expression levels following treatment with the STEMprep™ Mouse Tumor Dissociation Cocktail. Stable epitopes showed minimal or no change in detectability, moderate epitopes exhibited partial reduction in signal, and sensitive epitopes showed significant loss of expression. Over 200 epitopes were evaluated (Table 1), with the vast majority of cell surface epitopes (88.8%) being stable and detectable after incubation with the STEMprep™ Mouse Tumor Dissociation Cocktail.
Table 1. Assessment of Cell Surface Epitope Stability Following Treatment with the STEMprep™ Mouse Tumor Dissociation Cocktail
Figure 2. Majority of Cell Surface Epitopes Are Preserved After Incubation with the STEMprep™ Tumor Dissociation Cocktail
(A) Representative flow plots for each category: stable CD3 (clone 145-2C11), moderately affected CD371 (clone 5D3), and CD81 (clone Eat-2) that is sensitive to the enzyme cocktail. In the stable category, both staining intensity and the percent positive population remained unaffected by the STEMprep™ Mouse Tumor Dissociation Cocktail. The staining intensity was reduced in the moderate group, but the positive population was still distinct and similar to the untreated condition. Sensitive epitopes exhibited reduced detectability that may affect downstream experiments and conclusions. (B) Table summarizing the results of evaluating over 200 epitopes, with the vast majority of cell surface epitopes being stable and detectable after incubation with the STEMprep™ Mouse Tumor Dissociation Cocktail.
For epitopes sensitive to the STEMprep™ Mouse Tumor Dissociation Cocktail, staining optimization or use of alternative clones can be viable solutions. For more information, please contact us at techsupport@stemcell.com.
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