The Raji cell line is a human B-lymphocyte line derived from a patient with Burkitt lymphoma, and it has become one of the most widely used cell lines in immunology and cancer research worldwide.1PubMed Central. Integration of Epstein-Barr virus into chromosome 6q15 of Burkitt lymphoma cell line (Raji) induces loss of BACH2 expression Established in the 1960s, Raji cells carry several features that make them uniquely versatile: they harbor Epstein-Barr virus, display a characteristic chromosomal translocation involving the c-myc oncogene, and express a rich panel of B-cell surface markers. That combination has placed them at the center of research spanning antibody therapies, CAR-T cell development, virology, and drug discovery.
Where Raji Cells Came From
The Raji cell line was established from the tumor tissue of an 11-year-old Nigerian boy with Burkitt lymphoma, a fast-growing cancer of B lymphocytes that is especially common in equatorial Africa and strongly associated with Epstein-Barr virus (EBV) infection. The line was one of the earliest continuous human lymphoma cell lines to be successfully cultured, and it quickly became a reference standard for B-cell biology. Unlike many primary tumor samples that fail to grow indefinitely in culture, Raji cells proliferate vigorously in suspension, making them easy to maintain and scale up in the lab.
The Epstein-Barr Virus Connection
Raji cells are EBV-positive, carrying roughly 50 to 60 copies of the viral genome per cell.2Journal of Virology. Replication of latent Epstein-Barr virus genomes in Raji cells Most of those copies exist as circular DNA molecules floating freely in the nucleus rather than stitched into the cell’s own chromosomes. Density-transfer experiments showed that each of these viral plasmids replicates once per cell division, keeping the copy number stable over time.3Journal of Virology. Replication of latent Epstein-Barr virus genomes in Raji cells
EBV in Raji cells is predominantly latent, meaning the virus is not actively producing new infectious particles under normal culture conditions. However, the specific pattern of viral gene expression places Raji in a distinct latency category compared to other EBV-positive B-cell lines.4Journal of Virology. Chromatin Profiling of Epstein-Barr Virus Latency Control Region Researchers can chemically induce these cells to switch on the lytic (virus-producing) cycle, which has made Raji a go-to model for studying how EBV transitions between dormancy and active replication. One practical consequence: when Raji cells are chemically induced, they release exosomes containing the EBV-encoded dUTPase enzyme. Those exosomes can activate immune signaling in human dendritic cells and blood immune cells through a specific toll-like receptor, illustrating how a latently infected tumor cell can still shape the immune environment around it.5PLoS ONE. Epstein-Barr Virus Encoded dUTPase Containing Exosomes Modulate Innate and Adaptive Immune Responses in Human Dendritic Cells and Peripheral Blood Mononuclear Cells
The EBV status also has implications for how Raji cells compare to other Burkitt lymphoma lines. EBV-positive lines like Raji tend to have long telomeres, in the range of 10 to 19 kilobase pairs, while some EBV-positive lines carrying defective virus (like Daudi) have dramatically shorter telomeres of about 2.2 kilobase pairs.6PubMed. Telomere size and telomerase activity in Epstein-Barr virus (EBV)-positive and EBV-negative Burkitt’s lymphoma cell lines Those differences feed into broader questions about how EBV helps maintain chromosomal stability in infected cells.
Genetic Hallmarks
Like most Burkitt lymphoma cells, Raji carries a translocation between chromosomes 8 and 14, written as t(8;14). This rearrangement moves the c-myc oncogene from its normal home on chromosome 8 to a region on chromosome 14 that controls antibody heavy-chain production. Parked next to those powerful regulatory elements, c-myc is overexpressed, driving the relentless proliferation that defines the disease. In Raji specifically, c-myc lands in a switch region of the gamma heavy-chain locus.7PubMed Central. The translocated c-myc oncogene of Raji Burkitt lymphoma cells is not expressed in human lymphoblastoid cells An elegant set of fusion experiments showed that when Raji cells were hybridized with normal human lymphoblastoid cells, the translocated c-myc gene was silenced in the resulting hybrids, suggesting that normal cells possess regulatory mechanisms capable of shutting down the rogue oncogene.8PubMed Central. The translocated c-myc oncogene of Raji Burkitt lymphoma cells is not expressed in human lymphoblastoid cells
Beyond that signature translocation, Raji cells have a complicated karyotype. An analysis of 100 individual metaphase spreads identified four distinct subpopulations of cells, all carrying 48 chromosomes as the most common count. Across these cells, researchers catalogued 15 marker chromosomes, including the telltale 14q+ and 8q- markers that are hallmarks of the Burkitt translocation.9Cancer Genetics and Cytogenetics. Population analysis of karyotypic heterogeneity of the raji burkitt lymphoma cell line: Analysis of 100 karyotypes This heterogeneity is worth bearing in mind: any lab working with Raji cells is working with a mixture of closely related but genetically non-identical subclones, and the balance between them can drift with passage.
A separate consequence of the EBV genome in these cells became clear when researchers found that a portion of EBV DNA integrates directly into chromosome 6, disrupting the BACH2 gene, a transcription factor involved in normal B-cell development and tumor suppression.10PubMed Central. Integration of Epstein-Barr virus into chromosome 6q15 of Burkitt lymphoma cell line (Raji) induces loss of BACH2 expression This integration event is separate from the episomal EBV copies described earlier and represents a relatively uncommon accident that likely contributes to the biology of this particular cell line.
Surface Markers and the B-Cell Identity
Raji cells express a textbook set of B-cell surface proteins, which is a major reason they show up in so many experimental protocols. Flow cytometry consistently detects strong surface expression of CD19, CD20, and CD22, the trio of antigens most commonly targeted by antibody therapies and engineered T cells in B-cell cancers.11Frontiers in Genome Editing. CRISPR-mediated generation of a tumor-associated antigen-deficient Raji platform to investigate antigen loss in CAR-T cell therapy Among Burkitt lymphoma lines, Raji displays moderate to high CD20 levels; one comparison measured background-corrected mean fluorescence at 650 units for Raji versus 880 for Daudi and 400 for Ramos.12Clinical Cancer Research. Efficient Elimination of B-Lineage Lymphomas by Anti-CD20–Auristatin Conjugates
Another surface molecule of particular interest is CD21, also known as complement receptor 2 (CR2). CD21 binds fragments of the complement protein C3 that become attached to immune complexes and pathogens during complement activation.13European Journal of Immunology. CD21 augments antigen presentation in immune individuals This receptor doubles as the entry point for EBV itself, which is how Raji cells became infected in the first place. CD21 also turns out to be the gateway for a surprising interaction: HIV-1 can infect Raji cells even though these cells completely lack CD4, the receptor HIV normally uses. When HIV particles are coated with complement fragments (opsonized) in normal serum, they bind CD21 and enter the cell, producing a productive infection. Blocking CD21 with an antibody completely prevents this complement-mediated HIV entry.14Scandinavian Journal of Immunology. Complement Receptor Type 2 Mediates Infection of the Human CD4‐Negative Raji B‐Cell Line with Opsonized HIV This finding expanded the understanding of how HIV can reach cell types beyond its primary CD4-positive T-cell targets.
The Standard Platform for Antibody Killing Assays
If you have ever read a paper describing how well a therapeutic antibody kills lymphoma cells, there is a good chance Raji cells were the target. Two assay types dominate this space, and Raji serves as the workhorse for both.
In complement-dependent cytotoxicity (CDC) assays, an antibody like rituximab binds CD20 on the Raji cell surface, then recruits serum complement proteins that punch holes in the cell membrane. A typical protocol incubates rituximab at varying concentrations with Raji cells and rabbit complement, then measures how many cells lyse.15PubMed Central. Exploring complement-dependent cytotoxicity by rituximab isotypes in 2D and 3D-cultured B-cell lymphoma Among commonly tested lines, Raji is one of the most susceptible to CDC, with IgG1-form rituximab producing efficient killing.16PubMed Central. Exploring complement-dependent cytotoxicity by rituximab isotypes in 2D and 3D-cultured B-cell lymphoma This high sensitivity has made the Raji CDC assay a standard biopharmaceutical potency test used to evaluate biosimilar versions of rituximab and other anti-CD20 antibodies.17Scientific Reports. Label-free assessment of complement-dependent cytotoxicity of therapeutic antibodies via a whole-cell MALDI mass spectrometry bioassay
In antibody-dependent cellular cytotoxicity (ADCC) assays, rituximab-coated Raji cells serve as targets for natural killer (NK) cells. Researchers have used this system to show that genetic variants in the NK cell receptor that grabs the antibody’s tail end affect killing efficiency, an observation with real clinical relevance for predicting which patients respond best to rituximab.18PubMed Central. Effects of complement and serum IgG on rituximab-dependent natural killer cell-mediated cytotoxicity against Raji cells
CAR-T Cell Development
Chimeric antigen receptor (CAR) T-cell therapy has transformed the treatment of certain B-cell cancers, and Raji cells have been a central part of the testing pipeline from the earliest preclinical stages. Because Raji reliably expresses CD19, the primary target for most approved CAR-T products, it serves as the default positive-control target in co-culture killing assays. A typical experiment mixes anti-CD19 CAR-T cells with Raji target cells at various ratios to measure how efficiently the engineered T cells destroy the tumor line.19PubMed Central. Pre-clinical assessment of chimeric antigen receptor t cell therapy targeting CD19+ B cell malignancy
A newer line of research tackles one of the biggest clinical problems in CAR-T therapy: antigen loss. Tumors sometimes escape CAR-T cells by downregulating or losing the target antigen entirely. To model this, one group used CRISPR gene editing to create Raji sublines that individually lack CD19, CD20, or CD22 on their surface. Flow cytometry confirmed that these knockouts completely eliminated expression of each target while leaving the other markers intact.20Frontiers in Genome Editing. CRISPR-mediated generation of a tumor-associated antigen-deficient Raji platform to investigate antigen loss in CAR-T cell therapy These engineered Raji variants let researchers test whether dual-targeting or multi-targeting CAR designs can overcome antigen escape, a question that is critical for next-generation therapies.
Xenograft Models in Mice
Raji cells transplant readily into immunodeficient mice, creating tumor models that mimic disseminated lymphoma in a living system. The route of injection matters considerably. Intravenous inoculation produces the shortest survival, with tumors homing primarily to the liver. Subcutaneous injection gives the longest survival. Intraperitoneal injection leads to the widest spread, with the highest number of infiltrated organs and the most involvement of lymph node-bearing sites. All three routes achieve complete tumor take at inoculum sizes in the range of 100,000 to a million cells.21PubMed. Characterization of a xenograft model for anti-CD19 CAR T cell studies This flexibility allows researchers to pick a model that matches their experimental question: intravenous for an aggressive disseminated disease model, subcutaneous for easy tumor-size measurement, intraperitoneal for lymphatic spread.
Drug Discovery Beyond Antibodies
Raji cells are not limited to antibody testing. They also serve as a screening platform for small molecules and novel drug formats. An amonafide analogue called 7-b, for instance, was shown to inhibit Raji cell growth in a dose-dependent manner by arresting cells in the G1 phase of the cell cycle, generating reactive oxygen species, and triggering programmed cell death through the mitochondrial pathway.22PubMed. 7-b, a novel amonafide analogue, cause growth inhibition and apoptosis in Raji cells via a ROS-mediated mitochondrial pathway
A more unconventional approach uses drug-free macromolecular therapeutics. In this system, two polymer-based nanoconjugates each carry complementary nucleotide strands along with antibody fragments that recognize CD20. When both conjugates bind to CD20 molecules on Raji cells, the complementary strands hybridize, physically crosslinking the receptors without any traditional drug payload. This crosslinking alone triggers apoptosis. The system’s effectiveness depends on structural variables including the length of the complementary sequences, the number of pendant strands per polymer, and the polymer’s molecular weight, all of which were systematically optimized using Raji cells as the test bed.23PubMed Central. Drug-free macromolecular therapeutics: Impact of structure on induction of apoptosis in Raji B cells
Rituximab Resistance and What It Reveals
One limitation of using Raji cells as a static model is that tumors in patients evolve. To better mimic clinical drug resistance, researchers have generated rituximab-resistant Raji sublines (often abbreviated RRCL) by exposing cells to gradually increasing doses of the antibody over many passages. These resistant cells progressively lose CD20 from their surface, directly mirroring a mechanism of relapse seen in patients.24PubMed Central. Regulation of CD20 in rituximab-resistant cell lines and B-cell non-Hodgkin lymphoma The resistant sublines have been used to study how CD20 expression is regulated at the level of gene transcription and protein turnover, providing clues for strategies to re-sensitize resistant tumors.
Extracellular Vesicles as Biological Messengers
Raji cells shed extracellular vesicles (EVs), tiny membrane-bound particles that carry cargo including proteins, lipids, and small RNAs. These vesicles are not passive debris. When Raji-derived EVs are taken up by other lymphoma cells, they promote proliferation and invasion. The mechanism involves a specific microRNA, miR-106a, which the EVs deliver to recipient cells. That microRNA suppresses Beclin1, a protein involved in autophagy, effectively blocking the self-eating process that would otherwise slow tumor growth. The same effect was confirmed in mouse experiments, where Raji-derived EVs accelerated tumor growth in vivo.25PubMed Central. Lymphoma cell-derived extracellular vesicles inhibit autophagy and apoptosis to promote lymphoma cell growth via the microRNA-106a/Beclin1 axis
As noted in the EBV section, chemically induced Raji cells also release exosomes loaded with the EBV-encoded dUTPase enzyme, which activates innate immune pathways in dendritic cells and blood mononuclear cells. The dUTPase activity in exosomal fractions was roughly 700-fold enriched compared to the general culture supernatant, and about five-fold higher in exosomes from induced versus non-induced cells.26PLoS ONE. Epstein-Barr Virus Encoded dUTPase Containing Exosomes Modulate Innate and Adaptive Immune Responses in Human Dendritic Cells and Peripheral Blood Mononuclear Cells These two lines of research illustrate how Raji-derived vesicles can influence both tumor progression and the surrounding immune landscape.
Viral Permissiveness Beyond EBV
Raji cells are naturally resistant to many viruses, but researchers have exploited creative workarounds to use them as infection models. Dengue virus, for example, barely infects Raji cells even at high concentrations. But when all the components of a specific crosslinking cocktail involving dengue-antibody immune complexes and the B-cell receptor were present, infection became detectable, pointing to a B-cell receptor-dependent enhancement mechanism.27bioRxiv. B cell receptor dependent enhancement of dengue virus infection This approach uses Raji’s very resistance as an advantage: because baseline infection is nearly zero, any infection that does occur can be attributed cleanly to the specific entry mechanism being tested.
The complement-mediated HIV infection described earlier is another example of the same logic. Raji cells lack CD4 entirely, so any HIV entry must be happening through an alternative route, making the line a clean tool for isolating complement receptor-dependent viral entry.28Scandinavian Journal of Immunology. Complement Receptor Type 2 Mediates Infection of the Human CD4‐Negative Raji B‐Cell Line with Opsonized HIV
Practical Considerations for Lab Work
Raji cells grow in suspension, which simplifies many procedures but introduces its own challenges. Unlike adherent cells that can be inspected under a microscope for confluency, suspension cultures require cell counts to gauge density. The line grows aggressively and needs regular splitting to avoid overgrowth, which can alter surface marker expression and other phenotypic properties over time.
Transfecting Raji cells, introducing foreign DNA, was historically difficult because B-cell lines tend to resist chemical transfection methods. Electroporation solved this problem. Early work demonstrated that electroporation could achieve stable transformation frequencies suitable for establishing permanent transfectant lines from Raji and other lymphoid cells.29Nucleic Acids Research. Electroporation: application to human lympboid cell lines for stable introduction of a transactivator gene of human T-cell leukemia virus type I More recently, CRISPR-based gene editing has proven effective in Raji cells, as demonstrated by the clean antigen knockouts used in CAR-T escape modeling.30Frontiers in Genome Editing. CRISPR-mediated generation of a tumor-associated antigen-deficient Raji platform to investigate antigen loss in CAR-T cell therapy
Authentication is another practical concern. Because Raji is so widely used and has been passaged in thousands of laboratories over decades, cross-contamination with other cell lines is a real risk. Short tandem repeat (STR) profiling is the standard identity check, and Raji is included as a reference line in contamination-detection studies that use deep sequencing to catch even low-level mixing between cell lines.31Oxford Academic. Authentication, characterization and contamination detection of cell lines, xenografts and organoids by barcode deep NGS sequencing The karyotypic heterogeneity described earlier adds another layer of complexity: even an authenticated Raji culture can drift phenotypically if one subclone outcompetes the others. Labs that depend on specific marker expression levels, especially CD20 for rituximab assays, need to periodically verify those levels rather than assuming stability.
The EBV carried by Raji cells also raises biosafety considerations. Although the virus is latent under normal conditions and Raji cells carry a defective viral genome that does not efficiently produce infectious particles, labs still typically handle the line under Biosafety Level 2 conditions as a precaution against any residual reactivation risk.