Immortalization of shrimp lymphoid cells by hybridizing with the continuous cell line Sf9 leading to the development of 'PmLyO-Sf9 '.

Shrimp progressively gets more attention among marine invertebrates from researchers all over the world due to it being a healthy food as well as having economic importance. There were a lot of attempts to develop a continuous cell line from shrimp but none successful. In this context a novel hybrid cell line named 'PmLyO-Sf9' could be developed by fusing shrimp lymphoid organ cells with Sf9 cells after to metabolic blocking of Sf9 cells using puromycin and actinomycin D and effecting the fusion by way of PEG application. The cells are maintained and multiplied in a mixture of SCCM and TNM-FH having osmolality 550 mOsm kg-1 and pH 6.8. Transmission electron microscopy of the hybrid cells revealed the presence of two nuclei during the initial stages and a single nucleus subsequently. The cell line is with shrimp and Sf9 genomic components and shrimp specific protein and is susceptible to WSSV. Shrimp elongation factor, Sf9 beta-actin, shrimp STAT and peroxinectin could be expresses through RT-PCR in the cell line. This is the first successful report of a hybrid cell line with shrimp genomic components and envisaged to be recognized a model system for multitudes of biomedical research in vitro. The cell line is in the National Cell Line Repository of ICAR - National Bureaue of Fish Genetic Resources, Lucknow, India.

[At Home among Strangers: Is It Possible to Create Hypoimmunogenic Pluripotent Stem Cell Lines?]

Human pluripotent stem cells, which include embryonic stem cells and induced pluripotent cells (iPSCs), are capable of unlimited division and differentiation into all cells of the body. These cells are considered as a potential source of various types of cells for transplantations. The use of autologous iPSCs is not potentially associated with immune rejection and does not require immunosuppression required for allogeneic grafts. However, the high cost of this technology and the duration of obtaining iPSCs and differentiated cells may limit the use of autologous iPSCs in clinical practice. In addition, full equivalence and immunological compatibility of autologous iPSCs and their derivatives have been repeatedly questioned. One approach to solving the problem of the immunological compatibility of allogeneic derivatives of iPSCs can be the establishment of cell lines with reduced immunogenicity. Differentiated derivatives of such iPSCs may be suitable for transplantation to any patient. This review discusses the strategies for evading immune surveillance in normal and tumor processes that can be used to establish stem cell lines with reduced immunogenicity.

Stimulation of nuclear receptor REV-ERBs regulates tumor necrosis factor-induced expression of proinflammatory molecules in C6 astroglial cells.

Under physiological conditions, astrocytes maintain homeostasis in the CNS. Following inflammation and injury to the CNS, however, activated astrocytes produce neurotoxic molecules such as cytokines and chemokines, amplifying the initial molecular-cellular events evoked by inflammation and injury. Nuclear receptors REV-ERBα and REV-ERBß (REV-ERBs) are crucial in the regulation of inflammation- and metabolism-related gene transcription. The current study sought to elucidate a role of REV-ERBs in rat C6 astroglial cells on the expression of inflammatory molecules following stimulation with the neuroinflammatory cytokine tumor necrosis factor (TNF). Stimulation of C6 cells with TNF (10 ng/ml) significantly increased the mRNA expression of CCL2, interleukin-6 (IL-6), inducible nitric oxide synthase (iNOS), and matrix metalloprotease (MMP)-9, but not fibroblast growth factor-2 (FGF-2), cyclooxygenase-2 (COX-2) and MMP-2. Treatment with either REV-ERB agonists GSK4112 or SR9009 significantly blocked TNF-induced upregulation of CCL2 mRNA and MMP-9 mRNA, but not IL-6 mRNA and iNOS mRNA expression. Furthermore, treatment with RGFP966, a selective histone deacetylase 3 (HDAC3) inhibitor, potently reversed the inhibitory effects of GSK4112 on TNF-induced expression of MMP-9 mRNA, but not CCL2 mRNA. Expression of Rev-erbs mRNA in C6 astroglial cells, primary cultured rat cortical and spinal astrocytes was confirmed by reverse transcription polymerase chain reaction. Together, the findings demonstrate an anti-inflammatory effect, downregulating of MMP-9 and CCL2 transcription, of astroglial REV-ERBs activation through HDAC3-dependent and HDAC3-independent mechanisms.

A new human mast cell line expressing a functional IgE receptor converts to tumorigenic growth by KIT D816V transfection.

In systemic mastocytosis (SM), clinical problems arise from factor-independent proliferation of mast cells (MCs) and the increased release of mediators by MCs, but no human cell line model for studying MC activation in the context of SM is available. We have created a stable stem cell factor (SCF) -dependent human MC line, ROSA(KIT WT), expressing a fully functional immunoglobulin E (IgE) receptor. Transfection with KIT D816V converted ROSA(KIT WT) cells into an SCF-independent clone, ROSA(KIT D816V), which produced a mastocytosis-like disease in NSG mice. Although several signaling pathways were activated, ROSA(KIT D816V) did not exhibit an increased, but did exhibit a decreased responsiveness to IgE-dependent stimuli. Moreover, NSG mice bearing ROSA(KIT D816V)-derived tumors did not show mediator-related symptoms, and KIT D816V-positive MCs obtained from patients with SM did not show increased IgE-dependent histamine release or CD63 upregulation. Our data show that KIT D816V is a disease-propagating oncoprotein, but it does not activate MCs to release proinflammatory mediators, which may explain why mediator-related symptoms in SM occur preferentially in the context of a coexisting allergy. ROSA(KIT D816V) may provide a valuable tool for studying the pathogenesis of mastocytosis and should facilitate the development of novel drugs for treating SM patients.

Characterization of a resident population of adventitial macrophage progenitor cells in postnatal vasculature.

RATIONALE: Macrophages regulate blood vessel structure and function in health and disease. The origins of tissue macrophages are diverse, with evidence for local production and circulatory renewal. OBJECTIVE: We identified a vascular adventitial population containing macrophage progenitor cells and investigated their origins and fate. METHODS AND RESULTS: Single-cell disaggregates from adult C57BL/6 mice were prepared from different tissues and tested for their capacity to form hematopoietic colony-forming units. Aorta showed a unique predilection for generating macrophage colony-forming units. Aortic macrophage colony-forming unit progenitors coexpressed stem cell antigen-1 and CD45 and were adventitially located, where they were the predominant source of proliferating cells in the aortic wall. Aortic Sca-1(+)CD45(+) cells were transcriptionally and phenotypically distinct from neighboring cells lacking stem cell antigen-1 or CD45 and contained a proliferative (Ki67(+)) Lin(-)c-Kit(+)CD135(-)CD115(+)CX3CR1(+)Ly6C(+)CD11b(-) subpopulation, consistent with the immunophenotypic profile of macrophage progenitors. Adoptive transfer studies revealed that Sca-1(+)CD45(+) adventitial macrophage progenitor cells were not replenished via the circulation from bone marrow or spleen, nor was their prevalence diminished by depletion of monocytes or macrophages by liposomal clodronate treatment or genetic deficiency of macrophage colony-stimulating factor. Rather adventitial macrophage progenitor cells were upregulated in hyperlipidemic ApoE(-/-) and LDL-R(-/-) mice, with adventitial transfer experiments demonstrating their durable contribution to macrophage progeny particularly in the adventitia, and to a lesser extent the atheroma, of atherosclerotic carotid arteries. CONCLUSIONS: The discovery and characterization of resident vascular adventitial macrophage progenitor cells provides new insight into adventitial biology and its participation in atherosclerosis and provokes consideration of the broader existence of local macrophage progenitors in other tissues.

Lymphotoxin ß receptor activation on macrophages induces cross-tolerance to TLR4 and TLR9 ligands.

Our previous studies indicated that lymphotoxin ß receptor (LTßR) activation controls and downregulates inflammatory reactions. In this study, we report that LTßR activation on primary mouse macrophages results in induction of tripartite motif containing (TRIM) 30α, which negatively regulates NF-κB activation induced by TLR signaling. LTßR activation results in a downregulation of proinflammatory cytokine and mediator expression upon TLR restimulation, demonstrating that LTßR signaling is involved in the induction of TLR cross-tolerance. Specific knockdown experiments using TRIM30α-specific small interfering RNA abolished the LTßR-dependent induction of TRIM30α and LTßR-mediated TLR cross-tolerance. Concordantly, LTßR activation on bone marrow-derived macrophages induced cross-tolerance to TLR4 and TLR9 ligands in vitro. Furthermore, we have generated cell type-specific LTßR-deficient mice with ablation of LTßR expression on macrophages/neutrophils (LTßR(flox/flox) × LysM-Cre). In bone marrow-derived macrophages derived from these mice LTßR-induced cross-tolerance to TLR4 and TLR9 ligands was impaired. Additionally, mice with a conditional ablation of LTßR expression on macrophages (LTßR(flox/flox) × LysM-Cre) are resistant to LTßR-induced TLR4 tolerance in vivo. Collectively, our data indicate that LTßR activation on macrophages by T cell-derived lymphotoxin α(1)ß(2) controls proinflammatory responses by activation of a TRIM30α-controlled, counterregulatory signaling pathway to protect against exacerbating inflammatory reactions.

A germline-competent embryonic stem cell line from NOD.Cg-Prkdc ( scid ) Il2rg ( tm1Wjl )/SzJ (NSG) mice.

The NOD.Cg-Prkdc ( scid ) Il2rg ( tm1Wjl )/SzJ mouse strain, commonly known as NSG (for NOD SCID Gamma) is severely immunodeficient and thus is an excellent recipient for xenografts, and in particular for engrafting human tumor cells and human hematopoietic stem cells. In the latter case, these cells give rise to many human hematopoetic lineages in their NSG hosts, resulting in recapitulation of many of the features of a human immune system. However, the immune system of these "humanized mice" (huMice) is not completely functional, in part because of a lack of expression of necessary human cytokines and HLA molecules by NSG host tissues. In order to facilitate the genetic modification of this strain in order to improve the huMouse model, we have created germline competent ES cells of this strain in which such modifications can be carried out.

Characterization and virus susceptibility of a skin cell line from red-spotted grouper (Epinephelus akaara).

A red-spotted grouper Epinephelus akaara skin (RGS) cell line was established and characterized. RGS cells had a normal diploid chromosome number of 2n = 48, the morphology of which was fibroblastic-like in 3 days and epithelial-like over 5 after 16 passages. The cells multiplied well in Dulbecco's modified Eagle's medium supplemented with 10% of fetal bovine serum at 25°C. Susceptibilities of RGS and grass carp ovary (GCO) cells to two viruses were tested, and the results showed that the titer of an iridovirus Rana grylio virus (RGV) in RGS cells was 10³·5 TCID50 ml⁻¹, which was much higher than a rhabdovirus spring viremia of carp virus (SVCV) in the cells (10°·5 TCID50 ml⁻¹). The titers of RGV and SVCV in GCO were 106·° TCID50 ml⁻¹ and 108·° TCID50 ml⁻¹, respectively, which were higher than those in RGS cells. The data may imply that RGS cells could be selectively resistible to some viruses during infection. RT-PCR analysis of RGV-infected RGS cells showed that RGV could replicate in RGS cells. Further study of virus replications in RGS cells was conducted by electron microscopy and immunofluorescence microscopy has shown that virus particles scattered in the cytoplasm and virus protein appeared in both the cytoplasm and nucleus. The results suggested that RGS cells could be used as a potential in vitro model to study the cutaneous barrier function against virus infection.

[Comparative characteristics of new mesenchymal stem cell lines derived from human embryonic stem cells, bone marrow and foreskin].

New nonimmortalized fibroblast-like cell lines SC5-MSC and SC3a-MSC, FetMSC, FRSN were obtained from human embryonic stem cells (ESC), bone marrow of a 5-6-days embryo and foreskin of a 3-years-old boy, respectively. All the lines are successfully used as the feeder at human ESC cultivation. It is determined that the average cell population doublings time varies from 25.5 h for ISC5-MSC to 38.8 h for SC3a-MSC. Active proliferation of all the lines is also shown by the corresponding growth curves. Numerical and structural karyotypic analysis showed that these lines had normal karyotype: 46,XX (SC5-MSC and SC3a-MSC) and 46,XY (FetMSC and FRSN). To determine the status of the lines, their cell surface markers were analyzed by flow cytometry. This analysis revealed the presence of surface antigens CD44, CD73, CD90, CD105 and HLA-ABC, characteristic of human MSC, and the absence of CD34 and HLA-DR. Different lines were found to express CD117(c-kit) to a different level. Immunofluorescence and flow cytometry analysis did not detect TRA-1-60 and Oct-4, characteristic of human embryonic stem cells, and revealed interlinear variations in the level of SSEA, which did not depend on the cell origin. It is not clear yet whether these interlinear variations affect functional MSC status. In all the lines, immunofluorescence analysis showed the presence of the markers of early differentiation in the derivates of three germ layers which may allow MSC to be useful, in corresponding microenvironments, for reparation of tissue injures. Adipogenic and osteogenic differentiatiation of all cell lines has been shown.

Functional assessment of miR­1291 in colon cancer cells.

miR­1291 exerts an anti­tumor effect in a subset of human carcinomas, including pancreatic cancer. However, its role in colorectal cancer (CRC) is largely unknown. In the present study, the expression and effect of miR­1291 in CRC cells was investigated. It was identified that miR­1291 significantly suppressed the proliferation, invasion, cell mobility and colony formation of CRC cells. Additionally, miR­1291 induced cell apoptosis. A luciferase reporter assay revealed that miR­1291 directly bound the 3'­untranslated region sequence of doublecortin­like kinase 1 (DCLK1). miR­1291 also suppressed DCLK1 mRNA and protein expression in HCT116 cells that expressed DCLK1. Furthermore, miR­1291 suppressed cancer stem cell markers BMI1 and CD133, and inhibited sphere formation. The inhibitory effects on sphere formation, invasion and mobility in HCT116 cells were also explored and verified using DCLK1 siRNAs. Furthermore, miR­1291 induced CDK inhibitors p21WAF1/CIP1 and p27KIP1 in three CRC cell lines, and the overexpression of DCLK1 in HCT116 cells led to a decrease of p21WAF1/CIP1 and p27KIP1. Intravenous administration of miR­1291 loaded on the super carbonate apatite delivery system significantly inhibited tumor growth in the DLD­1 xenograft mouse model. Additionally, the resultant tumors exhibited significant upregulation of the p21WAF1/CIP1 and p27KIP1 protein with treatment of miR­1291. Taken together, the results indicated that miR­1291 served an anti­tumor effect by modulating multiple functions, including cancer stemness and cell cycle regulation. The current data suggested that miR­1291 may be a promising nucleic acid medicine against CRC.

DEAD/H BOX 3 (DDX3) helicase binds the RIG-I adaptor IPS-1 to up-regulate IFN-beta-inducing potential.

Retinoic acid-inducible gene-I (RIG-I)-like receptors (RLR) are members of the DEAD box helicases, and recognize viral RNA in the cytoplasm, leading to IFN-beta induction through the adaptor IFN-beta promoter stimulator-1 (IPS-1) (also known as Cardif, mitochondrial antiviral signaling protein or virus-induced signaling adaptor). Since uninfected cells usually harbor a trace of RIG-I, other RNA-binding proteins may participate in assembling viral RNA into the IPS-1 pathway during the initial response to infection. We searched for proteins coupling with human IPS-1 by yeast two-hybrid and identified another DEAD (Asp-Glu-Ala-Asp) box helicase, DDX3 (DEAD/H BOX 3). DDX3 can bind viral RNA to join it in the IPS-1 complex. Unlike RIG-I, DDX3 was constitutively expressed in cells, and some fraction of DDX3 is colocalized with IPS-1 around mitochondria. The 622-662 a.a DDX3 C-terminal region (DDX3-C) directly bound to the IPS-1 CARD-like domain, and the whole DDX3 protein also associated with RLR. By reporter assay, DDX3 helped IPS-1 up-regulate IFN-beta promoter activation and knockdown of DDX3 by siRNA resulted in reduced IFN-beta induction. This activity was conserved on the DDX3-C fragment. DDX3 only marginally enhanced IFN-beta promoter activation induced by transfected TANK-binding kinase 1 (TBK1) or I-kappa-B kinase-epsilon (IKKepsilon). Forced expression of DDX3 augmented virus-mediated IFN-beta induction and host cell protection against virus infection. Hence, DDX3 is an antiviral IPS-1 enhancer.

Some properties and applications of cell lines and clones established from tet-responsive-SV40 tag mice and mES cell lines.

In this article, 11 cell lines established from transgenic mice and mouse embryonic stem cells (mESC) expressing SV40Tag under control of tetracycline/doxycycline (tet-off, tet-on) are described. Several cell lines were further transfected with a plasmid vector containing genes coding for a cytokine/protein under tet-regulation to obtain tet-co-regulated expression of cytokine/protein. A total of 29 clones and 234 subclones have been established so far. Partial characterization of these tet-responsive cell lines, clones and subclones was performed. Questions related to the rare frequency of establishing permanent cycling cell lines from this source, the unusual expression pattern of SV40Tag protein in the subcellular compartment and the phenotype of 'stemness' of several such cell lines are raised. Some future applications of these cells, related to immunology and transplantation, are discussed.

Mapping the stem cell state: eight novel human embryonic stem and embryonal carcinoma cell antibodies.

The antigenic profile of human embryonic stem (ES) and embryonal carcinoma (EC) cells has served as a key element of their characterization, with a common panel of surface and intracellular markers now widely used. Such markers have been used to identify cells within the 'undifferentiated state', yet it appears that this categorization may be an oversimplification, because a number of sub-states appear to exist within this state. To increase the resolution of the undifferentiated state, we have generated eight novel monoclonal antibodies, all capable of recognizing undifferentiated human ES and EC cells, and herein describe their characterization. The reactivity of these antibodies against a range of cell lines is reported, as well as their developmental regulation, basic biochemistry and reactivity in immunohistochemistry of testicular germ cell tumours. Our data reveal a range of reactivity for all antibodies against both ES and EC cells, suggesting that these markers will afford recognition of unique sub-states within the undifferentiated stem cell compartment.

Cell surface immunoglobulin. 3. Isolation and characterization of immunoglobulin from nonsecretory human lymphoid cells.

Cells from an established line of Burkitt lymphoma (Daudi) were enzymatically radioiodinated, and labeled Ig from the cell surface was isolated and studied. Subcellular fractionation of labeled cells confirmed that intracellular proteins from the cytoplasm are not iodinated by this method. Radioactive Ig was identified as monomeric (8S) IgM, and an average of 10(5) Ig molecules was found per cell. Ig molecules could be released from the plasma membrane by detergent lysis under nonreducing conditions indicating that attachment of Ig to the plasma membrane occurs via noncovalent interactions. The ratio of micro/L radioactivity in surface Ig was the same as that of total cellular Ig radioiodinated in solution suggesting that a large portion of the Fc fragment is not buried within the membrane. In contrast to the results obtained with cell surface Ig, most intracellular Ig was found as "free" micro- and L chains regardless of whether lysates were labeled with (125)I or cells were labeled with leucine-(3)H. The results indicate that only a small percentage of the total Ig of Daudi cells is associated with the cell surface and suggest that covalent assembly of Ig occurs at or near the time that the molecule becomes part of the plasma membrane. Similarities between cell surface Ig on normal splenic lymphocytes and Daudi cells suggest that the latter is a neoplasm of bone marrow-derived lymphocytes.

Lack of distinctive surface antigen on cells transformed by murine sarcoma virus.

Some murine sarcoma virus (MSV)-transformed mouse 3T3 cells contain the MSV genome in the absence of infectious helper murine leukemia virus (MuLV) and MSV production. These cells, designated S+L- (sarcoma positive, leukemia negative), were analyzed for the presence of a possible MSV-determined membrane antigen by the mixed hemadsorption test and in vitro lymphocyte cytotoxicity assay. Two different serological approaches were used: (a) isoantibody-free sera were obtained by immunizing with MSV of syngeneic origin or by allowing primary, autologous MSV sarcomas to regress, or (b) alloantisera obtained by immunizing C57BL mice with S+L- cells were absorbed with the corresponding nontransformed 3T3 cells until all activity against 3T3 had been removed. While MuLV-superinfected S+L- cells and a culture line of an MSV sarcoma known to produce both MSV and MLV were highly reactive, normal 3T3 and S+L- cells were negative. Similarly, lymph node cells from MSV immune mice or rats did not kill S+L- cells, although they were cytotoxic against target cells known to carry MuLV-associated antigens. Thus, the present study gives no positive evidence for the existence of any MSV-induced new surface antigen in the transformed target cell, known to carry the viral genome.

Immunological functions of human T-lymphoid cell line (MOLT). I. Release of immunosuppressive factors from the mixture of MOLT-4 cells and sheep red blood cells.

A short term incubation of the mixture of established human T-lymphoid cells (MOLT) and sheep red blood cells (SRBC) resulted in the release of factors which nonspecifically suppressed the response of mouse spleen cells against heterologous erythrocytes in vitro. Neither human B-cell line (RPMI 1788), nor the supernate of MOLT cell suspension in the absence of SRBC had such suppressive effects. The supernate of the mixture of MOLT cells with chicken red blood cells (CRBC) did not suppress either anti-CRBC or anti-SRBC responses of mouse spleen cells. Since CRBC did not form rosettes with MOLT cells, it is suspected that the origin of the production of these factors might be MOLT cells forming SRBC rosettes. Some of these factors are dialysable.

Antigenic properties of murine sarcoma virus-transformed BALB-3T3 nonproducer cells.

The isolation of clonal lines of murine sarcoma virus-transformed, non-producer BALB/3T3 cells has provided a model system for determining whether RNA tumor virus-transformed cells possess virus-specific transplantation antigens. MSV nonproducer cells (K-234) were clonally derived from an inbred mouse cell line, BALB/3T3. A parallel virus-producing cell line was obtained by infection of the MSV nonproducer cells with Rauscher leukemia virus. K-234 was much more tumorigenic than K-234(R). Preimmunization of syngeneic mice with either K-234(R) or with UV-inactivated Rauscher leukemia virus induced transplantation resistance to subsequent challenge with K-234(R), but not with K-234. In contrast, mice preimmunized with nonproducer cells were not made resistant to subsequent challenge with the homologous cells. Antisera prepared from mice immunized with K-234(R) were specifically cytotoxic and positive by fluorescent antibody staining for K-234(R) target cells, but not to either BALB/3T3 or K-234. The results show that MSV nonproducer cells lack detectable transplantation antigens and suggest that the transplantation resistance to the producing cells is attributable to maturing virus at the cell surface.

Renal deposition of soluble immune complexes in mice bearing B-16 melanoma. Characterization of complexes and relationship to tumor progress.

Histologic and immunofluorescence studies of the kidneys of mice bearing a progressive melanoma show a proliferative glomerulonephritis associated with immune complex deposition in the mesangium and along the glomerular basement membrane This immune complex disease is distinct from the age-associated disease of the C57BL/6J host strain and the complexes can be shown to consist of soluble tumor antigen and antitumor antibody. Furthermore, the intensity of IgG complex deposition correlates directly with tumor progress (size and metastases) and inversely with mononuclear leukocyte infiltration of the tumor. In vitro assays for lymphocyte cytotoxicity and humoral antibody were found to be less reliable indicators of tumor progress. The possible role of circulating soluble tumor antigen in modifying the immune response to tumors is discussed.

Variation in susceptibility of a human lymphoid cell line to immune lysis during the cell cycle. Lack of correlation with antigen density and complement binding.

Cultured human lymphoid cells RPMI 8866 at different stages of their growth cycle vary in their susceptibility to lysis by rabbit, human, and guinea pig complement activated by HL-A antibodies or heterologous antibodies directed to membrane antigens; cells in G(1) phase are the least sensitive to lysis. To investigate the cause of differential susceptibility of cells RPMI 8866 to lysis, the expression of HL-A determinants and the ability of cells to react with complement were investigated. No change was detected in the density of HL-A antigens on RPMI 8866 cells in synchronous growth as determined by quantitative microabsorption assays, isotopic antiglobulin tests and yields of soluble HL-A antigens. Cells did not vary during the growth cycle in their ability to interact with complement components and in their capacity to activate the complement system through the classical or alternate pathway. These data suggest that variability in lytic susceptibility is due to changes in the structure of the cell membrane or in its ability to repair complement induced damage at certain intervals during the cell cycle. Therefore, this cell line constitutes a useful model to investigate the final steps of the cytolytic reaction.

Long-term establishment of a human plasmacyte cell line derived from a patient with IgD multiple myeloma. I. Requirement of a plasmacyte-stimulating factor for the proliferation of myeloma cells in tissue culture.

Cell line LA-49, derived from pleural fluid cells of a patient with IgD multiple myeloma, was established in culture and maintained for more than 1 yr. The D-myeloma protein produced in culture was similar to the serum D-myeloma protein in electrophoretic mobility and in delta- and lambda-chain antigens. The plasma cell tumor culture, LA-49, differed from numerous immunoglobulin-producing B-lymphoblastoid cell lines established in this laboratory in: (a) Morphology (revealing various stages of maturation); (b) type of immunoglobulin produced (IgD vs. IgM, IgG, and/or, rarely, IgA); (c) growth characteristics (requirement of plasmacyte-stimulating factor); and (d) chromosomal features (polyploid vs. pseudodiploid). A growth factor was needed for cell division and maintenance of culture viability. This factor was supplied readily by irradiated feeder layers of normal human fibroblasts or conditional media from fibroblast cultures. Preliminary characterization of this factor revealed it to be a protein with a mol wt of approximately 150,000 daltons.