t1b 196 cd4 cd70 b cells  (ATCC)

Journal: mAbs

Article Title: Insights into the molecular basis of a bispecific antibody's target selectivity

doi: 10.1080/19420862.2015.1022695

Figure Lengend Snippet: Schematic diagram showing conventional bivalent monospecific anti-CD4 and anti-CD70 IgGs along with anti-CD4/CD70 monovalent bispecific DuetMab.

Article Snippet: T1B-196 CD4 − /CD70 + B cells were obtained from the American Type Culture Collection and cultured in RPMI-1640 supplemented with 15% heat-inactivated (HI) fetal bovine serum (FBS).

Techniques:

Journal: mAbs

Article Title: Insights into the molecular basis of a bispecific antibody's target selectivity

doi: 10.1080/19420862.2015.1022695

Figure Lengend Snippet: Cell binding and ADCC activity of anti-CD4/CD70 DuetMab. ( A ) Anti-CD4/CD70 DuetMab exhibits preferential cell binding to CD4 + /CD70 + T cells via concurrent engagement to CD4 and CD70 on a single cell. ( B ) Anti-CD4/CD70 DuetMab preferentially kills CD4 + /CD70 + T cells as measured by ADCC. Each point represents the mean value of triplicate wells and the standard deviation is represented by error bars.

Article Snippet: T1B-196 CD4 − /CD70 + B cells were obtained from the American Type Culture Collection and cultured in RPMI-1640 supplemented with 15% heat-inactivated (HI) fetal bovine serum (FBS).

Techniques: Binding Assay, Activity Assay, Standard Deviation

Journal: mAbs

Article Title: Insights into the molecular basis of a bispecific antibody's target selectivity

doi: 10.1080/19420862.2015.1022695

Figure Lengend Snippet: CD4 and CD70 receptor density on human lymphocytes

Article Snippet: T1B-196 CD4 − /CD70 + B cells were obtained from the American Type Culture Collection and cultured in RPMI-1640 supplemented with 15% heat-inactivated (HI) fetal bovine serum (FBS).

Techniques:

Journal: mAbs

Article Title: Insights into the molecular basis of a bispecific antibody's target selectivity

doi: 10.1080/19420862.2015.1022695

Figure Lengend Snippet: Binding affinity of IgG and DuetMab to CD4 and CD70

Article Snippet: T1B-196 CD4 − /CD70 + B cells were obtained from the American Type Culture Collection and cultured in RPMI-1640 supplemented with 15% heat-inactivated (HI) fetal bovine serum (FBS).

Techniques: Binding Assay

Journal: mAbs

Article Title: Insights into the molecular basis of a bispecific antibody's target selectivity

doi: 10.1080/19420862.2015.1022695

Figure Lengend Snippet: Cell binding of various DuetMabs variants. Binding of anti-CD4/CD70 DuetMab variants to ( A ) CD4 + /CD70 + , ( B ) CD4 − /CD70 + and ( C ) CD4 + /CD70 − lymphocytes in a mixture of all 3 cell types. All variants with reduced affinity to CD4 exhibited improved binding selectivity over the parental DuetMab whereas their binding to target CD4 + /CD70 + T cells was not substantially impaired. Each point represents the mean values of triplicate wells and the standard deviation is represented by error bars.

Article Snippet: T1B-196 CD4 − /CD70 + B cells were obtained from the American Type Culture Collection and cultured in RPMI-1640 supplemented with 15% heat-inactivated (HI) fetal bovine serum (FBS).

Techniques: Binding Assay, Standard Deviation

Journal: mAbs

Article Title: Insights into the molecular basis of a bispecific antibody's target selectivity

doi: 10.1080/19420862.2015.1022695

Figure Lengend Snippet: ADCC activity of various DuetMabs variants. ( A ) Selective ADCC depletion of CD4 + /CD70 + T cells in a cell-mixture also containing non-target CD4 + /CD70 − T cells and CD4 − /CD70 + B cells. ( B ) ADCC activity of parental and anti-CD4 VκY94A+V H Y99A/CD70 DuetMabs against individual populations of CD4 + /CD70 + and CD4 + /CD70 − T-lymphocytes at varying E:T ratios. Each point in these studies represents the mean values of triplicate wells and the standard deviation is represented by error bars.

Article Snippet: T1B-196 CD4 − /CD70 + B cells were obtained from the American Type Culture Collection and cultured in RPMI-1640 supplemented with 15% heat-inactivated (HI) fetal bovine serum (FBS).

Techniques: Activity Assay, Standard Deviation

Journal: mAbs

Article Title: Insights into the molecular basis of a bispecific antibody's target selectivity

doi: 10.1080/19420862.2015.1022695

Figure Lengend Snippet: Cytotoxicity of various CD4 affinity-reduced DuetMab variants

Article Snippet: T1B-196 CD4 − /CD70 + B cells were obtained from the American Type Culture Collection and cultured in RPMI-1640 supplemented with 15% heat-inactivated (HI) fetal bovine serum (FBS).

Techniques:

Journal: mAbs

Article Title: Insights into the molecular basis of a bispecific antibody's target selectivity

doi: 10.1080/19420862.2015.1022695

Figure Lengend Snippet: Effect of antibody valence on cell binding and ADCC activity. ( A ) Cell binding and ( B ) ADCC activity of anti-CD4 VκY94A+V H Y99A/CD70 and 2 monospecific (anti-CD4 VκY94A+V H Y99A/NMGC and anti-CD70/NMGC) DuetMabs at equimolar concentration against CD4 + /CD70 + T cells alone. ( C ) Non-target ADCC activity of anti-CD4 variants formatted as either monovalent anti-CD4/CD70 DuetMab or bivalent anti-CD4 IgG against CD4 + /CD70 − T cells alone. Each point in these studies represents the mean values of triplicate wells and the standard deviation is represented by error bars.

Article Snippet: T1B-196 CD4 − /CD70 + B cells were obtained from the American Type Culture Collection and cultured in RPMI-1640 supplemented with 15% heat-inactivated (HI) fetal bovine serum (FBS).

Techniques: Binding Assay, Activity Assay, Concentration Assay, Standard Deviation

Journal: British Journal of Cancer

Article Title: Melanoma-expressed CD70 is involved in invasion and metastasis

doi: 10.1038/bjc.2015.412

Figure Lengend Snippet: CD70 ectopic expression in human melanomas. Representative images of CD70 expression after staining by IHC paraffin-embedded melanoma human biopsies ( A ) or staining of cryostat sections of frozen melanoma tumour specimens ( B ). Negative staining corresponded to CD70 expression in <2% of melanoma cells, moderate staining in 2–20% and strong staining in more than 20%. ( C ) Three slides of the same melanoma fragment showing that CD70 expression is in the tumour cells. The slides were labelled with anti-CD70 mAb, anti-MelanA or anti-KBA.62 antitumour antibodies. ( D ) Primary human melanomas contain more CD70+ tumour cells than metastases as quantified in paraffin-embedded melanoma biopsies. ( E ) The intensity of CD70 expression is also stronger in primary melanomas than in metastases. P -values were evaluated using the Mann–Whitney test ( D ) or the χ 2 -test ( E ). ** P <0.005; *** P <0.001.

Article Snippet: B16F10-CD70 and its control cell line B16F10-wt were produced in our laboratory ( Cormary et al , 2005 ) by genetic modification of B16F10 murine melanoma cells purchased from the ATCC.

Techniques: Expressing, Staining, Negative Staining, MANN-WHITNEY

Journal: British Journal of Cancer

Article Title: Melanoma-expressed CD70 is involved in invasion and metastasis

doi: 10.1038/bjc.2015.412

Figure Lengend Snippet: CD70 expression decreases with time and disease progression in human melanomas. ( A ) CD70 membrane expression measured by flow cytometry in three representative melanoma cell lines: LB1319-MEL; LB39-MEL; and LB39-MEL clones CD70+ vs CD70−. ( B ) Decrease of membrane CD70 expression in three melanoma cell lines derived from the same patient: LB33-MEL.A (skin, 1988); LB33-MEL.B (lymph node, 1993); and LB33-MEL.D (intestine, 1999). ( C ) Decrease with disease progression of melanoma-expressed CD70: primary tumours, skin and lymph node metastases from the same patient C121084. ( D ) Monomeric (CD70 M) and trimeric (CD70 T) forms of CD70 detected by western blot in LB1319-MEL (total, cytosolic and membrane fractions), B16F10-wt and B16F10-CD70 cells.

Article Snippet: B16F10-CD70 and its control cell line B16F10-wt were produced in our laboratory ( Cormary et al , 2005 ) by genetic modification of B16F10 murine melanoma cells purchased from the ATCC.

Techniques: Expressing, Flow Cytometry, Clone Assay, Derivative Assay, Western Blot

Journal: British Journal of Cancer

Article Title: Melanoma-expressed CD70 is involved in invasion and metastasis

doi: 10.1038/bjc.2015.412

Figure Lengend Snippet: CD70 expression is associated with decreased in vivo metastatic capacity. ( A ) C57BL/6, C57BL/6 IFN- γ KO and NMRI nu/nu mice were injected i.v. with B16F10-wt (blue) or B16F10-CD70 (red) cells and pulmonary metastases were quantified, showing that B16F10-CD70 cells induced less metastases implantations. ( B ) Representative lung photomicrographs (left) and enlargement of metastatic areas (right) of C57BL/6 mice injected i.v. with B16F10-wt (top) or B16F10-CD70 (bottom) cells. Arrows indicate metastases. P -values were evaluated using the t -test. *** P <0.001.

Article Snippet: B16F10-CD70 and its control cell line B16F10-wt were produced in our laboratory ( Cormary et al , 2005 ) by genetic modification of B16F10 murine melanoma cells purchased from the ATCC.

Techniques: Expressing, In Vivo, Injection

Journal: British Journal of Cancer

Article Title: Melanoma-expressed CD70 is involved in invasion and metastasis

doi: 10.1038/bjc.2015.412

Figure Lengend Snippet: In melanoma cells CD70 expression is associated with decreased in vitro migration and invasion capacities. Decreased in vitro migration ( A ) and invasion ( B ) capacities of B16F10-CD70+ cells analysed using, respectively, transwells and invasion chambers with B16F10-wt vs B16F10-CD70 cells. In the upper chamber the medium was or not (NT) supplemented with Ctrl Ig (Ct) or anti-CD70 mAb (QA). Decreased in vitro migration capacity of B16F10-CD70+ cells was analysed by wound-healing experiments done with B16F10-wt and B16F10-CD70 cells ( C ). Decreased in vitro migration ( D ) and invasion ( E ) capacities of CD70+ human cells: LB39-MEL CD70− clone (blue), LB39-MEL CD70+ clone (red) and LB1319-MEL cells (black). Results ( A , B , D and E ) are expressed as mean values ( n =7). P -values were evaluated using the Tukey ANOVA test. * P <0.05; ** P <0.005; *** P <0.001.

Article Snippet: B16F10-CD70 and its control cell line B16F10-wt were produced in our laboratory ( Cormary et al , 2005 ) by genetic modification of B16F10 murine melanoma cells purchased from the ATCC.

Techniques: Expressing, In Vitro, Migration

Journal: British Journal of Cancer

Article Title: Melanoma-expressed CD70 is involved in invasion and metastasis

doi: 10.1038/bjc.2015.412

Figure Lengend Snippet: CD70 signalling modulates MAPK activation, RhoE and ROCK1 expression, and cytoskeleton formation. Western blot analysis ( A ) and quantification ( B ) in arbitrary units (a.u.) of CD70 M decrease and CD70 T increase in LB1319-MEL cells incubated with anti-CD70 mAb (QA). Controls are untreated (NT) and Ctrl Ig (Ct)-incubated LB1319-MEL cells. ( C ) Western blots analyses of the MEK/ERK pathway activation and RhoE overexpression in these LB1319-MEL cells. ( D ) The 3D invasion using LB1319-MEL spheroids treated with the Abs plus MEK inhibitor U0126, showing anti-CD70-induced invasion enhancement and U0126-induced invasion inhibition. ( E ) Western blot quantification of total ROCK1 and ROCK2 in LB1319-MEL cells incubated with Ctrl or anti-CD70 mAb, showing anti-CD70-induced ROCK1 inhibition. ( F ) Disappearance of stress fibres (actin) and focal adhesions (vinculin) in LB1319-MEL cells treated with anti-CD70 mAb for 48 h. Results in B and E are expressed as mean values ( n =3). P -values were evaluated using the Tukey ANOVA test ( B ) or t -test ( E ). ** P <0.005.

Article Snippet: B16F10-CD70 and its control cell line B16F10-wt were produced in our laboratory ( Cormary et al , 2005 ) by genetic modification of B16F10 murine melanoma cells purchased from the ATCC.

Techniques: Activation Assay, Expressing, Western Blot, Incubation, Over Expression, Inhibition

Journal: Cell Death & Disease

Article Title: CD70-restricted specific activation of TRAILR1 or TRAILR2 using scFv-targeted TRAIL mutants

doi: 10.1038/cddis.2013.555

Figure Lengend Snippet: CD70-specific mAb l α hCD70 is a potent inhibitor of CD27–CD70 interaction. ( a ) OVCAR3 and Mino cells were preincubated (1 h) with the indicated concentrations of the CD70-specific mAbs l α hCD70 and 1F6. Cells were then incubated with 200 ng/ml CD27-Fc-GpL for an additional hour and after removal of unbound molecules cell-associated GpL activity was determined. ( b ) Mino and U266 cells were preincubated (1 h) with the indicated concentrations of the CD70-specific mAbs l α hCD70 and 1F6 and were then used to stimulate IL8 production in HT1080-CD27 cells that were cultured in a 96-well plate. Next day, cell culture supernatants were analyzed for the presence of IL8 by ELISA. ( c ) Monomeric and trimeric GpL fusion proteins of a l α hCD70-derived scFv were used for equilibrium binding studies with OVCAR3 and Mino cells

Article Snippet: To obtain l α hCD70 (27B3), a Fab fragment phage library derived from llamas immunized with 786-O cells (ATCC) was screened for antibodies interfering with CD70-CD27 interaction and high affinity for CD70 essentially as described elsewhere.

Techniques: Incubation, Activity Assay, Cell Culture, Enzyme-linked Immunosorbent Assay, Derivative Assay, Binding Assay

Journal: Cell Death & Disease

Article Title: CD70-restricted specific activation of TRAILR1 or TRAILR2 using scFv-targeted TRAIL mutants

doi: 10.1038/cddis.2013.555

Figure Lengend Snippet: Initial characterization of scFv:CD70-TRAIL fusion proteins. ( a ) Scheme of scFv:l α hCD70-TRAIL fusion proteins. scFv:l α hCD70, CD70-specific scFv; F, Flag tag; TNC, tenascin-C trimerization domain; TRAIL, TRAILmutR1 and TRAILmutR2, aa 95–281 of wild-type TRAIL and TRAILR1- and TRAILR2-specific mutants derived thereof. ( b ) SDS-PAGE analysis of purified scFv:l α hCD70-TNC-TRAIL fusion proteins (100 ng each). ( c ) HT1080 cells were transiently transfected with a CD70 expression construct or empty vector. Next day, transfected cells were stimulated for an additional day with the indicated concentrations of scFv:l α hCD70-TNC-TRAIL, scFv:l α hCD70-TNC-TRAILmutR1 and scFv:l α hCD70-TNC-TRAILmutR2 in the presence and absence of a competing conventional CD70-specific antibody

Article Snippet: To obtain l α hCD70 (27B3), a Fab fragment phage library derived from llamas immunized with 786-O cells (ATCC) was screened for antibodies interfering with CD70-CD27 interaction and high affinity for CD70 essentially as described elsewhere.

Techniques: FLAG-tag, Derivative Assay, SDS Page, Purification, Transfection, Expressing, Construct, Plasmid Preparation

Journal: Cell Death & Disease

Article Title: CD70-restricted specific activation of TRAILR1 or TRAILR2 using scFv-targeted TRAIL mutants

doi: 10.1038/cddis.2013.555

Figure Lengend Snippet: CD70-restricted apoptosis induction by scFv:l α hCD70-TRAIL, scFv:l α hCD70-TRAILmutR1 and scFv:l α hCD70-TRAILmutR2. ( a ) The indicated cell lines were analyzed by using FACS for CD70 cell surface expression. ( b ) OVCAR3, Mino and Jurkat cells were cultured in 96-well plates, and half of the cells were pretreated with 10 μ g/ml of a scFv:l α hCD70 competing conventional CD70-specific antibody. Cells were then challenged overnight in triplicates with the indicated concentrations of scFv:l α hCD70-TNC-TRAIL, scFv:l α hCD70-TNC-TRAILmutR1 and scFv:l α hCD70-TNC-TRAILmutR2. Finally, cellular viability was determined using the MTT assay or crystal violet staining. OVCAR3 cells were challenged in the presence of 2.5 μ g/ml CHX, which sensitizes this cell line for apoptosis induction. ( c ) The indicated cell line cells were stimulated in the presence and absence of the conventional CD70-specific antibody l α hCD70 for 4–6 h with 100 ng/ml of scFv:l α hCD70-TNC-TRAIL, scFv:l α hCD70-TNC-TRAILmutR1 and scFv:l α hCD70-TNC-TRAILmutR2. Total cell lysates were analyzed by western blotting for processing of the indicated proteins

Article Snippet: To obtain l α hCD70 (27B3), a Fab fragment phage library derived from llamas immunized with 786-O cells (ATCC) was screened for antibodies interfering with CD70-CD27 interaction and high affinity for CD70 essentially as described elsewhere.

Techniques: Expressing, Cell Culture, MTT Assay, Staining, Western Blot

Journal: Cell Death & Disease

Article Title: CD70-restricted specific activation of TRAILR1 or TRAILR2 using scFv-targeted TRAIL mutants

doi: 10.1038/cddis.2013.555

Figure Lengend Snippet: Blockade of CD70 has no effect on cell death induction by conventional TRAIL. ( a ) Mino cells were cultured in 96-well plates, and half of the cells were pretreated with 5 μ g/ml of scFv:l α hCD70-TNC-GpL and then challenged overnight in triplicates with the indicated concentrations of TNC-TRAIL, TNC-TRAILmutR1 and TRAILmutR2 with and without anti-Flag (1 μ g/ml) oligomerization. Cellular viability was determined using the MTT assay. ( b ) FACS analysis of CD70 and CD27 expression of Raji and KMS12.BM cells. ( c ) Raji and KMS12.BM cells were cultured in 96-well plates in the presence and absence of 10 μ g/ml of a scFv:l α hCD70 competing conventional CD70-specific antibody and were stimulated overnight in triplicates with the indicated concentrations of the various scFv:l α hCD70-TRAIL fusion proteins. Cellular viability was determined using the MTT assay

Article Snippet: To obtain l α hCD70 (27B3), a Fab fragment phage library derived from llamas immunized with 786-O cells (ATCC) was screened for antibodies interfering with CD70-CD27 interaction and high affinity for CD70 essentially as described elsewhere.

Techniques: Cell Culture, MTT Assay, Expressing

Journal: Cell Death & Disease

Article Title: CD70-restricted specific activation of TRAILR1 or TRAILR2 using scFv-targeted TRAIL mutants

doi: 10.1038/cddis.2013.555

Figure Lengend Snippet: Cell surface expression of CD27 and CD70 in cancer cell lines

Article Snippet: To obtain l α hCD70 (27B3), a Fab fragment phage library derived from llamas immunized with 786-O cells (ATCC) was screened for antibodies interfering with CD70-CD27 interaction and high affinity for CD70 essentially as described elsewhere.

Techniques: Expressing

Journal: Cell Death & Disease

Article Title: CD70-restricted specific activation of TRAILR1 or TRAILR2 using scFv-targeted TRAIL mutants

doi: 10.1038/cddis.2013.555

Figure Lengend Snippet: CD70-specific mAb l α hCD70 is a potent inhibitor of CD27–CD70 interaction. ( a ) OVCAR3 and Mino cells were preincubated (1 h) with the indicated concentrations of the CD70-specific mAbs l α hCD70 and 1F6. Cells were then incubated with 200 ng/ml CD27-Fc-GpL for an additional hour and after removal of unbound molecules cell-associated GpL activity was determined. ( b ) Mino and U266 cells were preincubated (1 h) with the indicated concentrations of the CD70-specific mAbs l α hCD70 and 1F6 and were then used to stimulate IL8 production in HT1080-CD27 cells that were cultured in a 96-well plate. Next day, cell culture supernatants were analyzed for the presence of IL8 by ELISA. ( c ) Monomeric and trimeric GpL fusion proteins of a l α hCD70-derived scFv were used for equilibrium binding studies with OVCAR3 and Mino cells

Article Snippet: To obtain l α hCD70 (27B3), a Fab fragment phage library derived from llamas immunized with 786-O cells (ATCC) was screened for antibodies interfering with CD70-CD27 interaction and high affinity for CD70 essentially as described elsewhere.

Techniques: Incubation, Activity Assay, Cell Culture, Enzyme-linked Immunosorbent Assay, Derivative Assay, Binding Assay

Journal: Cell Death & Disease

Article Title: CD70-restricted specific activation of TRAILR1 or TRAILR2 using scFv-targeted TRAIL mutants

doi: 10.1038/cddis.2013.555

Figure Lengend Snippet: Initial characterization of scFv:CD70-TRAIL fusion proteins. ( a ) Scheme of scFv:l α hCD70-TRAIL fusion proteins. scFv:l α hCD70, CD70-specific scFv; F, Flag tag; TNC, tenascin-C trimerization domain; TRAIL, TRAILmutR1 and TRAILmutR2, aa 95–281 of wild-type TRAIL and TRAILR1- and TRAILR2-specific mutants derived thereof. ( b ) SDS-PAGE analysis of purified scFv:l α hCD70-TNC-TRAIL fusion proteins (100 ng each). ( c ) HT1080 cells were transiently transfected with a CD70 expression construct or empty vector. Next day, transfected cells were stimulated for an additional day with the indicated concentrations of scFv:l α hCD70-TNC-TRAIL, scFv:l α hCD70-TNC-TRAILmutR1 and scFv:l α hCD70-TNC-TRAILmutR2 in the presence and absence of a competing conventional CD70-specific antibody

Article Snippet: To obtain l α hCD70 (27B3), a Fab fragment phage library derived from llamas immunized with 786-O cells (ATCC) was screened for antibodies interfering with CD70-CD27 interaction and high affinity for CD70 essentially as described elsewhere.

Techniques: FLAG-tag, Derivative Assay, SDS Page, Purification, Transfection, Expressing, Construct, Plasmid Preparation

Journal: Cell Death & Disease

Article Title: CD70-restricted specific activation of TRAILR1 or TRAILR2 using scFv-targeted TRAIL mutants

doi: 10.1038/cddis.2013.555

Figure Lengend Snippet: CD70-restricted apoptosis induction by scFv:l α hCD70-TRAIL, scFv:l α hCD70-TRAILmutR1 and scFv:l α hCD70-TRAILmutR2. ( a ) The indicated cell lines were analyzed by using FACS for CD70 cell surface expression. ( b ) OVCAR3, Mino and Jurkat cells were cultured in 96-well plates, and half of the cells were pretreated with 10 μ g/ml of a scFv:l α hCD70 competing conventional CD70-specific antibody. Cells were then challenged overnight in triplicates with the indicated concentrations of scFv:l α hCD70-TNC-TRAIL, scFv:l α hCD70-TNC-TRAILmutR1 and scFv:l α hCD70-TNC-TRAILmutR2. Finally, cellular viability was determined using the MTT assay or crystal violet staining. OVCAR3 cells were challenged in the presence of 2.5 μ g/ml CHX, which sensitizes this cell line for apoptosis induction. ( c ) The indicated cell line cells were stimulated in the presence and absence of the conventional CD70-specific antibody l α hCD70 for 4–6 h with 100 ng/ml of scFv:l α hCD70-TNC-TRAIL, scFv:l α hCD70-TNC-TRAILmutR1 and scFv:l α hCD70-TNC-TRAILmutR2. Total cell lysates were analyzed by western blotting for processing of the indicated proteins

Article Snippet: To obtain l α hCD70 (27B3), a Fab fragment phage library derived from llamas immunized with 786-O cells (ATCC) was screened for antibodies interfering with CD70-CD27 interaction and high affinity for CD70 essentially as described elsewhere.

Techniques: Expressing, Cell Culture, MTT Assay, Staining, Western Blot

Journal: Cell Death & Disease

Article Title: CD70-restricted specific activation of TRAILR1 or TRAILR2 using scFv-targeted TRAIL mutants

doi: 10.1038/cddis.2013.555

Figure Lengend Snippet: Blockade of CD70 has no effect on cell death induction by conventional TRAIL. ( a ) Mino cells were cultured in 96-well plates, and half of the cells were pretreated with 5 μ g/ml of scFv:l α hCD70-TNC-GpL and then challenged overnight in triplicates with the indicated concentrations of TNC-TRAIL, TNC-TRAILmutR1 and TRAILmutR2 with and without anti-Flag (1 μ g/ml) oligomerization. Cellular viability was determined using the MTT assay. ( b ) FACS analysis of CD70 and CD27 expression of Raji and KMS12.BM cells. ( c ) Raji and KMS12.BM cells were cultured in 96-well plates in the presence and absence of 10 μ g/ml of a scFv:l α hCD70 competing conventional CD70-specific antibody and were stimulated overnight in triplicates with the indicated concentrations of the various scFv:l α hCD70-TRAIL fusion proteins. Cellular viability was determined using the MTT assay

Article Snippet: To obtain l α hCD70 (27B3), a Fab fragment phage library derived from llamas immunized with 786-O cells (ATCC) was screened for antibodies interfering with CD70-CD27 interaction and high affinity for CD70 essentially as described elsewhere.

Techniques: Cell Culture, MTT Assay, Expressing