Written and maintained by CASRAI Editorial Board
Last updated
Short answer: a cell surface marker is a molecule, usually a protein or carbohydrate, exposed on the outer surface of a cell and detectable with a binding reagent such as an antibody. Researchers read several markers together, not one, to say what kind of cell they are looking at. For human leukocyte molecules the shared naming system is the CD (cluster of differentiation) nomenclature.
Definition and scope
A surface marker is defined by where it sits and how it is detected: it is on the outside of the plasma membrane, so an antibody can reach it on intact cells. That is why surface markers are the standard handle for sorting and phenotyping live populations by flow cytometry. Results are written with plus and minus signs (for example CD45+ or CD34-) to record whether a population carries the marker. The phrase cell surface markers in the plural usually means exactly this: the set of such molecules measured on a sample.
How markers are named: the CD system
The Human Cell Differentiation Molecules (HCDM) organisation describes its mission as characterising the structure, function and distribution of leucocyte surface molecules. Antibodies that show similar reaction patterns are grouped into a cluster of differentiation, and the cluster receives a CD number. HCDM lists characterised molecules from CD1 to CD350, and the first of the Human Leukocyte Differentiation Antigen (HLDA) workshops was held in Paris in 1982. Workshops rely on multi-laboratory blind analysis of antibodies.
Two points are easy to miss. First, a CD name is allocated when there is a need for a name, so not every molecule on an immune cell surface has one. Second, the order has reversed over time: molecules are now usually identified first by genetics or proteomics, and antibodies are produced afterwards. Single-marker pages on this site, such as CD45, CD11b and CD68, cover individual molecules; this page covers the shared logic.
Worked example: why a panel, not one marker
The ISCT position statement of Dominici and colleagues (Cytotherapy 2006;8(4):315-317) defines minimal criteria for multipotent mesenchymal stromal cells. As summarised in a 2023 scoping review (Fonseca et al., Heliyon 2023;9(2):e13464), the criteria combine four things:
| Criterion | What it requires |
|---|---|
| Adherence | Adherence to plastic under standard culture conditions |
| Positive surface markers | Expression of CD105, CD90 and CD73 |
| Negative surface markers | Absence of the haematopoietic markers CD45, CD34, CD14 or CD11b, CD79a or CD19, and HLA-DR |
| Differentiation | In vitro differentiation into osteoblasts, adipocytes and chondrocytes |
The lesson is structural. No single marker in the table identifies the cell type: the identity call comes from the positive and negative markers together, plus functional evidence. The negative markers do as much work as the positive ones, because they exclude blood-lineage cells.
Controls that make a marker call trustworthy
The flow cytometry guidelines by Cossarizza and colleagues (European Journal of Immunology 2021;51:2708-3145, third edition) group controls into three categories: setup (instrument) controls, specificity controls used for gating, and biological controls. Points from the specificity section that bear directly on surface markers:
- FMO control. The guidelines state that a fluorescence-minus-one control identifies spectral overlap of fluorochromes into the channel of interest. They also note that an FMO control cannot account for non-specific binding of the antibody it replaces; for that, use a biological control or another cell type in the sample.
- Isotype control. The guidelines say an isotype control is not useful for determining or excluding non-specific binding, and that isotype-matched gating controls have largely been superseded by FMO controls.
- Fc blocking. Non-specific binding can be reduced with blocking agents (Fc blockers).
- Viability stain. Viability stains are described as essential when analysing viable samples, because dead cells bind antibodies non-specifically and are more autofluorescent.
A checklist built from these points: confirm that the call rests on a marker set rather than a single marker; that boundaries were set with FMO or biological controls rather than an isotype-matched gating control; that Fc receptors were blocked where relevant; and that dead cells were excluded. Sample preparation affects all of this, as described in the PBMC preparation and staining guide, and sorting by markers is covered in cell sorting: nozzle, pressure, purity and viability.
Common misunderstandings
A marker is not an identity
A marker records presence or absence of one molecule. The ISCT criteria above require a pattern of positive and negative markers plus functional tests, not a single readout.
A CD number is not a quality grade
A CD number records that antibodies grouped into a cluster recognise a defined molecule. It says nothing about the performance of any particular antibody product, which is why specificity controls are still needed.
Isotype controls do not settle positivity
Many protocols still include them, but the Cossarizza guidelines say an isotype control is not useful for determining or excluding non-specific binding, and that isotype-matched gating controls have largely been superseded by FMO controls.
FAQ
What is the difference between a cell surface marker and a CD marker?
A cell surface marker is any detectable molecule on the outside of a cell. A CD marker is a surface marker that has been given a cluster-of-differentiation number through the HCDM process. Not every surface molecule has a CD name.
How many markers define a cell population?
There is no fixed number. The ISCT mesenchymal stromal cell criteria use three positive markers and several negative ones, together with adherence and differentiation tests.
Sources
- Human Cell Differentiation Molecules (HCDM), hcdm.org, read 2026-11-08.
- Cossarizza A et al. Guidelines for the use of flow cytometry and cell sorting in immunological studies (third edition). Eur J Immunol 2021;51:2708-3145. doi:10.1002/eji.202170126. PMC11115438, read 2026-11-08.
- Dominici M et al. Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement. Cytotherapy 2006;8(4):315-317.
- Fonseca LN et al. Cell surface markers for mesenchymal stem cells related to the skeletal system: a scoping review. Heliyon 2023;9(2):e13464. doi:10.1016/j.heliyon.2023.e13464. PMC9970931, read 2026-11-08.
General laboratory information only; it is not a protocol or a substitute for the instructions of the antibody or instrument manufacturer.
