Correct option is D
The correct answer is (d), which involves using DAPI for nucleic staining, CD19-FITC and CD3-PE for identifying T cells, and both CD8-CY7 PE and CD4-Cy7 PE for distinguishing between CD8+ and CD4+ T cells.
In this experiment, the goal is to accurately identify and sort the CD8+ and CD4+ T cell populations. Here's the reasoning:
DAPI is a nuclear stain, which helps in identifying and excluding dead cells. It will allow only live cells (those with intact nuclei) to be analyzed.
CD19-FITC targets B lymphocytes. This is useful for distinguishing B cells, but in this case, it will help remove unwanted cell populations from the analysis, ensuring you focus on T cells.
CD3-PE is used to mark all T cells (both CD8+ and CD4+), as CD3 is a pan-T cell marker.
CD8-CY7 PE and CD4-Cy7 PE are used to differentiate CD8+ (cytotoxic) T cells from CD4+ (helper) T cells. These markers bind specifically to the CD8 and CD4 proteins, which are essential for distinguishing these two subsets of T cells.
This sequence of fluorochrome conjugates allows for the simultaneous detection and sorting of the CD8+ and CD4+ T cell populations, ensuring accurate results.
Additional Information
(a) DAPI and CD3-PE: While CD3-PE is used for T cell identification, this option does not include the markers required to distinguish between CD8+ and CD4+ T cells. Without the CD8-CY7 PE and CD4-Cy7 PE markers, this combination will not effectively separate these two important T cell subsets.
(b) DAPI; CD3-PE; CD8-CY7 PE or CD4-Cy7 PE: This option lacks the inclusion of CD19-FITC, which is useful for excluding B cells. While the use of DAPI and CD3-PE helps identify live T cells, the absence of CD19-FITC could result in unwanted B cell contamination in the analysis, especially when dealing with mixed cell populations.
(c) CD19-FITC and CD3-PE; CD8-CY7 PE or CD4-Cy7 PE: While it includes CD19-FITC for B cell identification, the combination does not involve DAPI, which is critical for live cell analysis. Additionally, CD19-FITC is unnecessary for sorting T cells and adds extra complexity to the process without contributing to the specific analysis of CD8+ and CD4+ T cells.
