Multicolor flowcytometric immunophenotyping is a valuable tool for detection of intraocular lymphoma

Ophthalmology. 2013 May;120(5):991-6. doi: 10.1016/j.ophtha.2012.11.007. Epub 2013 Feb 4.


Objective: Intraocular lymphoma (IOL) is a rare condition and frequently difficult to distinguish from uveitis or other uveitis-masquerading syndromes. The diagnosis is confirmed by cytologic examination of ocular fluid specimens and more recently by molecular-immunoglobulin heavy chain (IGH) translocation or cytokine analysis. However, some of these more recent methods have not been validated by follow-up studies.

Design: Evaluation of a diagnostic test.

Participants: In a cohort of 51 consecutive patients with a clinical suspicion of IOL, vitreous analysis was performed via multicolor flowcytometric immunophenotyping.

Methods: Multicolor flowcytometric immunophenotyping was performed with CD45, CD3, CD19, CD20, anti-SmIgκ, and anti-SmIgλ antibodies. The presence of a clear B-cell population showing a disequilibrium of Igκ versus Igλ expression was used to confirm the diagnosis of non-Hodgkin lymphoma (NHL). Patients were followed for a minimum of 2 years (mean, 5.9 ± 2.0 years) to validate the accuracy of the method.

Main outcome measures: The presence or absence of IOL during follow-up.

Results: In 14 of 51 patients, a clinical diagnosis of IOL was confirmed using flowcytometric analysis. Of these 14 patients, 11 had primary IOL and 3 had metastasized secondary lymphomas. In 3 of 51 patients who were diagnosed with (central nervous system) NHL during follow-up, the test failed to confirm the presence of a clonal B-cell population. In 18 of the 34 other patients, an infectious or well-defined immunologic disorder was established during follow-up. The remaining 16 patients, with a minimal follow-up of 2 years, were diagnosed with idiopathic uveitis.

Conclusions: Multicolor flowcytometric analysis had 82.4% sensitivity and 100% specificity in patients with suspected IOL. This is comparable to the reported vitreous interleukin (IL)-6/IL-10 testing sensitivity of 0.8 and sensitivity of 0.65 to 0.95 by immunoglobulin heavy chain (IGH) gene arrangement testing in clinical cohorts. Because flowcytometric tests are readily performed in hematologic laboratories, this can be regarded as a useful method for confirming the clinical diagnosis of IOL.

Financial disclosure(s): The author(s) have no proprietary or commercial interest in any materials discussed in this article.

MeSH terms

  • Aged
  • Antigens, CD / analysis
  • B-Lymphocytes / immunology
  • Cohort Studies
  • Eye Neoplasms / diagnosis*
  • Eye Neoplasms / immunology
  • Female
  • Flow Cytometry / methods*
  • Gene Rearrangement / genetics
  • Humans
  • Immunoglobulin Heavy Chains / genetics
  • Immunoglobulin Heavy Chains / metabolism
  • Immunophenotyping / methods*
  • Lymphoma, Non-Hodgkin / diagnosis*
  • Lymphoma, Non-Hodgkin / immunology
  • Male
  • Middle Aged
  • Rare Diseases / diagnosis*
  • Rare Diseases / immunology
  • Sensitivity and Specificity


  • Antigens, CD
  • Immunoglobulin Heavy Chains