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        Murine No Compensation Panels

Murine No Compensation Panels

Murine peripheral blood panels

Immunophenotype without Compensation

The no compensation panels below have been optimized for murine peripheral blood and secondary lymphoid tissue such as spleen, bone marrow and lymph nodes, to help identify common cell subsets. Useful to create a simple four color panel or a great starting point when designing larger and more complex panels.These panels have been optimized by titrating the antibody to give the best signal resolution.

General identification panels
To identify T, B, NK and myeloid cells.
Lymphocyte panels
To identify T and B cell populations such as naïve, memory and activated as well as helper and cytotoxic T cells.
NK cell and myeloid panels
To identify NK and NKT cells and various myeloid populations.

General Identification Panels

T, B and myeloid cells

Markers	Formats	Catalog #	Dilution
CD3	Pacific Blue	MCA500PB	Neat
CD19	Alexa Fluor 647	MCA1439A647	1:10
CD11b	RPE-Alexa Fluor 750	MCA74P750	1:10
Ly6G	FITC	MCA6077F	1:10

Cells	Surface Markers
T cells	CD3⁺ CD11b^-CD19^-Ly6G^-
B cells	CD3^- CD11b^- CD19⁺ Ly6G^-
Monocytes/Macrophages	CD3^- CD11b⁺ CD19^- Ly6G^-
Granulocytes	CD3^- CD11b⁺ CD19^- Ly6G⁺

Fig 1. General panel. A, Identification of T and B cells. B, Identification of myeloid cells from the CD3- CD19- population. Data acquired on the ZE5 Cell Analyzer.

Fig. 1. General panel. A, identification of T and B cells. B, identification of myeloid cells from the CD3^- CD19^- population. Data acquired on the ZE5^TM Cell Analyzer.

Isotype controls - Reagents to determine specific binding

T, B, NK and myeloid cells

Markers	Formats	Catalog #	Dilution
CD3	Pacific Blue	MCA500PB	Neat
B220	RPE-Alexa Fluor 750	MCA1258P750	1:10
CD49b	FITC	MCA2125F	1:5
CD11b	Alexa Fluor 647	MCA74A647	1:10

Cells	Surface Markers
T cells	CD3⁺ CD19^- CD49^- CD11b^-
B cells	CD3^- CD19⁺ CD49^- CD11b^-
NK cells	CD3^- CD19^- CD49b⁺ CD11b^-/+
Myeloid	CD3^- CD19^- CD49b^- CD11b⁺

Fig. 1. General panel.

Fig. 2. General panel. A, identification of T and B cells. B, identification of NK cells and myeloid cells using CD3, B220, CD49b and CD11b. Data acquired on the ZE5 Cell Analyzer.

Lymphocyte Panels

T cells

Markers	Formats	Catalog #	Dilution
CD3	Pacific Blue	MCA500PB	Neat
CD8	RPE-Alexa Fluor 750	MCA609P750	1:5
CD4	Alexa Fluor 647	MCA4635A647	1:2
CD45	FITC	MCA1031F	1:5

Cells	Surface Markers
T helper	CD45⁺ CD3⁺ CD4⁺ CD8^-
T cytotoxic	CD45⁺ CD3⁺ CD8⁺ CD4^-

Fig. 2. T cell panel.

Fig. 3. T cell panel. A, identification of T cells. B, identification of T helper and T cytotoxic cells using CD45, CD3, CD4 and CD8. Data acquired on the ZE5 Cell Analyzer.

Incorporating CD44 and CD62L into this panel will allow identification of naïve, memory and effector T cells for both the helper and cytotoxic populations.

Cells	Surface Markers
Naive T helper cells	CD3⁺ CD4⁺ CD62L⁺ CD44^-
Memory T helper cells	CD3⁺ CD4⁺ CD62L^-CD44⁺
Effector T helper cells	CD3⁺ CD4⁺ CD62L^- CD44^mid
Naïve T cytotoxic cells	CD3⁺ CD8⁺ CD62L⁺ CD44^-
Memory T cytotoxic cells	D3⁺ CD8⁺ CD62L^- CD44⁺
Effector T cytotoxic cells	CD3⁺ CD8⁺ CD62L^-CD44^mid

Activated T cells

Markers	Formats	Catalog #	Dilution
CD3	Pacific Blue	MCA500PB	Neat
CD4 or CD8	Alexa Fluor 647	MCA4635A647 or MCA609A647	1:2 or 1:5
B220	RPE-Alexa Fluor 750	MCA1258P750	1:10
CD69	FITC	MCA1580F	1:2

Cells	Surface Markers
Resting T helper cells	CD3⁺ B220^- CD4⁺ CD69^-
Activated T helper cells	CD3⁺ B220^- CD4⁺ CD69⁺
Resting T cytotoxic cells	CD3⁺ B220^- CD8⁺ CD69^-
Activated T cytotoxic cells	CD3⁺ B220^- CD8⁺ CD69⁺

B cells

Markers	Formats	Catalog #	Dilution
CD19	Pacific Blue	MCA1439PB	1:10
B220	RPE-Alexa Fluor 750	MCA1258P750	1:10
IgM	FITC
CD23	Alexa Fluor 647

Cells	Surface Markers
Immature B cells	CD19⁺ B220⁺ IgM⁺CD23^-
Mature B cells	CD19⁺ B220⁺ IgM^- CD23⁺

B cell maturation in the blood and bone marrow can also be defined using IgM and IgD with immature B cells IgM ⁺ IgD^- and mature B cells IgM^-IgD ⁺ . Other markers can be used such as CD93 and CD21.

Naïve and memory B cells

Markers	Formats	Catalog #	Dilution
CD19	PB	MCA1439PB	1:10
B220	RPE-Alexa Fluor 750	MCA1258P750	1:10
CD23	Alexa Fluor 647	MCA1511FT	Neat
CD80	Alexa Fluor 647	MCA2462A647	1:5

Cells	Surface Markers
Naïve B cells	CD19⁺ B220⁺ CD23⁺ CD80^-
Memory B cells	CD19⁺ B220⁺ CD23^- CD80⁺

The panel above is a simplification of the staining used to identify naïve and memory B cells, as the expression patterns of these surface markers can be variable compared to CD27 ⁺ and IgD expression in humans, which are more defined markers for naïve and memory B cells. Whilst this panel gives an indication of the amount of naïve and memory B cells, other markers could be included for more conclusive identification of these cells and other populations such as marginal zone B cells, depending upon the tissue being stained. These markers include CD38, CD62L, CD73, CD21 and PD-L2.

NK cell and Myeloid Panels

NK and NKT cells

Markers	Formats	Catalog #	Dilution
CD3	Pacific Blue	MCA500PB	Neat
CD335	RPE-Cy7	MCA5938PECy7	Neat
CD45	Alexa Fluor 647	MCA1031A647	1:10
CD122	FITC	MCA2363F	1:10

Cells	Surface Markers
NK precursors	CD45⁺ CD3^- CD122⁺ CD335^-
NK cells	CD45⁺ CD3^- CD122⁺ CD335⁺
NKT cells	CD45⁺ CD3⁺ CD122⁺ CD335⁺

Fig 4. NK, NKT and NK precursor cells

Fig 4. NK, NKT and NK precursor cells. A, identification of NK lineage cells using CD122. B, identification of CD335⁺ NK and CD335 ⁺ CD3 ⁺ NKT cells in blood. C, CD122 ⁺ CD335^- NK cell precursors can also be seen in bone marrow. Data acquired on the ZE5 Cell Analyzer.

Myeloid Cell Panel

Markers	Formats	Catalog #	Dilution
CD11b	Pacific Blue	MCA711PB	1:10
CD11c	RPE-Alexa Fluor 750	MCA1369P750	1:2
Ly6C	Alexa Fluor 647	MCA2389A647	1:10
Ly6G	FITC	MCA6077F	1:10

Cells	Surface Markers
Granulocytes	CD3^- CD11b⁺CD11c^-Ly6G⁺
Classical Monocytes	Ly6C^hi CD11b⁺CD11c^-Ly6G^-
Non-Classical monocytes	Ly6C^lo CD11b⁺CD11c^-Ly6G^-
Dendritic cells	CD11c⁺CD11b⁺Ly6C^+/- Ly6G^+/-

Fig 5. Myeloid cells

Fig 5. Myeloid cells. A, identification of dendritic cells (DC) and granulocytes. B, monocytes. C, Classical Ly6C^hi and non-classical Ly6C^lo monocytes. Data acquired on the ZE5 Cell Analyzer.

Although these no compensation panels are not available as pre-mixed four color panels cocktails, they have been optimized on the ZE5 Cell Analyzer and titrated for optimal resolution. To purchase pre-mixed panels for human, mouse, rat or dog cell surface markers, visit our antibody panels and cocktails webpage. If you can’t find the antibody you are interested in, you can search our database of over 3000 flow cytometry tested antibodies. To help you analyze your samples, information on the proportion of these populations in murine blood and tissues can be found on our cell frequency page.

When performing multicolor flow cytometry, you should consider careful sample preparation, Fc blocking, inclusion of a live/dead dye, doublet exclusion and using the appropriate controls. For more information, watch our flow cytometry webinar Optimize your Flow Cytometry or visit our application resources page.

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