Flow cytometry and the CD4 count !
- Topic
Flow cytometry counts and sorts cells one by one with lasers and fluorescent antibodies. It is how the CD4 count in HIV is measured, for example.
Updated 2026-11-01
What it is
In flow cytometry, cells pass in single file past a laser; the light they scatter reports their size and internal complexity, and monoclonal antibodies tagged with fluorochromes reveal which CD markers they carry. Gating separates populations such as CD4 T cells, CD8 T cells, B cells and NK cells. It is central to following HIV, classifying leukemias and lymphomas and studying immunodeficiencies.
Step by step
- Blood is stained with fluorescent antibodies against CD45, CD3, CD4, CD8 and others.
- Cells pass one at a time in front of the laser.
- Forward scatter estimates size; side scatter, granularity.
- Detectors record the fluorescence of each marker.
- Lymphocytes are gated (bright CD45, low granularity).
- Subsets are counted and, with reference beads, reported as an absolute count per microlitre.
Markers
- CD45 (all leukocytes)
- CD3 (T cells)
- CD4 (helper T)
- CD8 (cytotoxic T)
- CD19 and CD20 (B cells)
- CD16 and CD56 (NK)
- CD34 (progenitors)
Clinical notes
- Immunophenotyping of leukemias and lymphomas: lineage, maturation and clonality.
- Immunodeficiencies: absent CD19 B cells in X-linked agammaglobulinemia; very low T cells in severe combined immunodeficiency.
- Paroxysmal nocturnal hemoglobinuria: loss of CD55 and CD59 on red cells and granulocytes.
- CD34 cell counts for hematopoietic progenitor transplantation.
In the lab
- Normal adult CD4 count: about 500 to 1200 cells per mm³.
- In HIV, fewer than 200 CD4 cells per mm³ defines AIDS; in young children the CD4 percentage is used.
- CD4 measures immune damage; viral load measures viral replication. They are read together.
- Internal check: T, B and NK cells should add up to close to 100 % of lymphocytes.
Test yourself
Think of the answer, then tap the question to see it.
What do forward and side scatter tell you in flow cytometry?
Forward scatter relates to cell size and side scatter to granularity or internal complexity.
Which CD4 count defines AIDS in a person with HIV?
Fewer than 200 cells per mm³ (or an AIDS-defining illness).
Which cytometry finding suggests X-linked agammaglobulinemia?
Absent or near-absent B cells (CD19 or CD20) in peripheral blood.
Which markers are lost in paroxysmal nocturnal hemoglobinuria?
CD55 and CD59, GPI-anchored proteins that protect cells from complement.
Related
- ELISA, western blot and rapid tests
- Live vaccines: who should not receive them
- CD markers: the table to know
- CD4⁺ helper T cell
- B-cell (antibody) deficiencies: XLA, CVID, IgA deficiency, hyper-IgM
- Secondary immunodeficiency: HIV, malnutrition, drugs, asplenia
Related conditions
Microorganisms
Educational content on the immune system; not a treatment or individual vaccination guide and no substitute for a health professional's advice. No doses are given.
Sources: Immunophenotyping of Peripheral Blood and Bone Marrow Cells by Flow Cytometry (EJIFCC (PubMed Central), 2006); CD4 Lymphocyte Count (MedlinePlus (U.S. National Library of Medicine), 2024).
Spotted a mistake?We are always improving for the community. Tell us what is wrong and our team will review it.Report a mistake
Representative model, for educational purposes only.