biotinylated recombinant human cd19  (ACROBiosystems)

Journal: Cell Reports Medicine

Article Title: Engineered T cells stimulate dendritic cell recruitment and antigen spreading for potent anti-tumor immunity

doi: 10.1016/j.xcrm.2025.102307

Figure Lengend Snippet: FX-armed CAR-T cells demonstrate superior tumor control in both murine and humanized mouse models (A) Schematic depicting the constructs of mouse B7-H3 and CD19 CARs. (B–D) Average tumor growth curves of B16F10-CD19 (B) and B16F10-B7-H3 (C); survival curves of B16F10-B7-H3 tumor-bearing mice (D) ( n = 5 mice). (E and F) Absolute number of CAR-T cells and endogenous CD8 + T cells in B16F10-CD19 (E) and B16F10-B7-H3 (F) tumors. (G) Average tumor growth curves of the B16F10-B7-H3 model ( n = 8–10 mice). (H) Schematic depicting the humanized B7-H3 CAR constructs. (I and J) Average tumor growth curves of the PC3 tumor model (I) and survival curves of the A375-B7-H3 tumor model in Flt3ko&hFLT3LG NSG mice (J). (K) Schematic of the humanized Flt3KO&hFLT3LG NSG mice model. (L–N) Average tumor growth curves of A375-B7-H3 (L) and PC3 (M). Survival curves of PC3 tumor-bearing Flt3ko&hFLT3LG NSG mice (N). All data represent the mean ± SEM and were analyzed by Student’s t test for (E), (F), and (K) or two-way ANOVA with Tukey’s multiple-comparisons test for (B), (C), (G), (I), (L), and (M) or log rank (Mantel-Cox) test for (D), (J), and (N). Statistical difference is delineated by ns, not significant, ∗ p < 0.05 and ∗∗ p < 0.01.

Article Snippet: Biotinylated Recombinant Human CD19 Protein , Yeasen , Cat#94100ES20.

Techniques: Control, Construct

Journal: Science signaling

Article Title: Generation of antitumor chimeric antigen receptors incorporating T cell signaling motifs.

doi: 10.1126/scisignal.adp8569

Figure Lengend Snippet: Fig. 3. CD19–CAR-TK con- structs show robust efficacy in an in vivo NSG leukemia model. (A) Schematic of the NSG mouse model of leuke- mia. ADT, adoptive transfer. (B) Leukemia growth was evaluated at the indicated times by bioluminescent im- aging (BLI), and IVIS images are shown. GL, GFP-luciferase. (C) Quantification of the BLI radiance data. Data are means ± SEM. Statistical differences were assessed with a Mann- Whitney U test to compare CAR-TK2 (P values in purple) or CAR-TK3 (P values in blue) with the 28ζ-CAR. *P ≤ 0.05 and **P ≤ 0.01. (D to F) Flow cytometry analysis of periph- eral blood on day 30 (D30) assessing CD3+ cells (D), CAR+ cells (E), and T cell subsets as follows: T naïve (Tn; CD62L+

Article Snippet: Solid- phase plate stimulation Pierce streptavidin- coated high- capacity 96- well plates (Thermo Fisher Scientific, catalog no. 15500) were washed three times with PBS, and dilutions of biotinylated CD19 (R&D Systems, catalog no. AVI 9269) in PBS were added to each well in 100 μl and incubated for 60 min at 37°C.

Techniques: In Vivo, Adoptive Transfer Assay, Luciferase, MANN-WHITNEY, Flow Cytometry

Journal: Nature Communications

Article Title: Regulatory T cells expressing CD19-targeted chimeric antigen receptor restore homeostasis in Systemic Lupus Erythematosus

doi: 10.1038/s41467-024-46448-9

Figure Lengend Snippet: A Fox19CAR lentiviral vector (LV) and FoxP3 LV schematic representations. The FoxP3 gene and an anti-CD19 second-generation CAR are inserted in an unidirectional LV and under the control of a Phosphoglycerate Kinase (PGK) promoter. The vector encoding for the FoxP3 gene alone is a bi-directional LV with the FoxP3 gene in sense (PGK promoter) and the enhanced Green Fluorescent Protein (eGFP) in antisense (minimal CMV or mCMV promoter). LTR long terminal repeat, SD splice donor, SA splice acceptor, GA gag-pol element, RRE REV responsive element, cPPT central polypurine tract, pA polyadenilation signal, CTE constitutive transport element, WPRE woodchuck hepatitis virus post-transcriptional regulatory element. B Transduction efficiency of CAR19.28z LV, Fox19CAR LV and FoxP3 LV measured as percentage of either CAR + or GFP + cells among CD4 + CD25 + CD127 - FoxP3 + lymphocytes at day +14. Recombinant CD19 for UT-, 19CAR- and Fox19CAR-Tregs and GFP for FoxP3 LV respectively, were employed to assess the transduction percentage. N = 5 for UT- and 19CAR-Tregs. N = 10 for Fox19CAR-Tregs. N = 3 for FoxP3-Tregs. One-way ANOVA with Tukey correction for multiple comparison. *** p -value 0.0008, **** p -value < 0.0001. C Representative flow cytometry plot for co-localization of the CAR construct and FoxP3 in Fox19CAR-Tregs compared to untransduced (UT) cells. CAR expression was assessed using the human rCD19. D Expansion rate of UT-, 19CAR-, Fox19CAR- and FoxP3-Tregs, assessed at day +14 since the initial stimulation. N = 6 for UT-, Fox19CAR- and 19CAR-Tregs. N = 3 for FoxP3-Tregs. One-way ANOVA with Tukey correction for multiple comparison. E Polyclonal suppressive capacities of UT-, 19CAR- and Fox19CAR-Tregs. Results are expressed as Suppression Index, calculated as Suppression index = [1-(PBMCs’ proliferation with Tregs)/(PBMCs’ proliferation alone)] * 100. N = 3 for UT-, Fox19CAR- and 19CAR-Tregs. N = 3 for UT conventional T cells. Two-way ANOVA with Tukey correction for multiple comparison. F Antigen-specific suppressive capacities of UT-, 19CAR-, Fox19CAR- and FoxP3-Tregs. Results are expressed as Suppression index = [1-(B cell proliferation with Tregs)/(B cell proliferation alone)] * 100. N = 6 for UT- and Fox19CAR-Tregs. N = 3 for 19CAR- and FoxP3-Tregs. Two-way ANOVA with Tukey correction for multiple comparison. The exact p -values are reported in the graph. All the results are expressed as mean ± standard deviation.

Article Snippet: Fox19CAR engineered cells were identified using biotinylated recombinant human CD19 reagent (Miltenyi, 130-129-550, 1:67).

Techniques: Plasmid Preparation, Control, Virus, Transduction, Recombinant, Comparison, Flow Cytometry, Construct, Expressing, Standard Deviation

Journal: Nature Communications

Article Title: Regulatory T cells expressing CD19-targeted chimeric antigen receptor restore homeostasis in Systemic Lupus Erythematosus

doi: 10.1038/s41467-024-46448-9

Figure Lengend Snippet: A Generation of the humanized mouse model of SLE. 1 day-old pups of NSG mice were irradiated and transplanted with 0.8-1 × 10 5 human cord-blood stem cells/mouse injected intra-liver. The engraftment was monitored assessing the presence and the composition of human leukocytes on peripheral blood weekly by flow cytometry. After the establishment of a human immune system, mice were injected i.p. with pristane to induce a chronic inflammation. After 3 weeks, UT-, Fox19CAR-Tregs or PBS were injected and their kinetic and the levels of human leukocytes in the peripheral blood were monitored weekly by flow cytometry. Mice were sacrificed 15 weeks after humanization. B , C Longitudinal assessment of human B and T lymphocytes in peripheral blood. Human B and T cells were identified as huCD45 + CD19 + and huCD45 + CD3 + lymphocytes, respectively. Absolute cell counts were assessed by flow cytometry. Pristane injection is indicated with an arrow. N = 53 mice. One-way ANOVA with Tukey correction for multiple comparison. * p -value 0.021, **** p -value < 0.0001. D Longitudinal assessment of circulating Fox19CAR-Tregs in mouse peripheral blood after their injection. CAR + cells were identified with FITC-conjugated recombinant CD19 (rCD19). CAR-Treg absolute counts were assessed by flow cytometry. N = 16 mice. E – G Absolute numbers of circulating human cells in humanized mice in the different groups of treatment after the injection of Fox19CAR-Tregs, UT-Tregs or PBS. Human cells were identified as total huCD45 + leukocytes, huCD45 + CD3 + T and huCD45 + CD19 + B lymphocytes, respectively. Absolute cell counts were assessed by flow cytometry. N = 31 (16 CAR-Tregs, 11 UT-Tregs, 4 PBS). Two-way ANOVA test with Tukey correction for multiple comparisons. The exact p -values are reported in the graphs. All the results are expressed as mean ± standard deviation.

Article Snippet: Fox19CAR engineered cells were identified using biotinylated recombinant human CD19 reagent (Miltenyi, 130-129-550, 1:67).

Techniques: Irradiation, Injection, Flow Cytometry, Comparison, Recombinant, Standard Deviation

Journal: Nature Communications

Article Title: Regulatory T cells expressing CD19-targeted chimeric antigen receptor restore homeostasis in Systemic Lupus Erythematosus

doi: 10.1038/s41467-024-46448-9

Figure Lengend Snippet: A Percentage of regulatory T cells in the different organs at sacrifice in the different groups of treatment. Tregs were defined as CD3 + CD4 + CD25 + CD127 - FoxP3 + lymphocytes and their frequency was assessed by flow cytometry. The results are expressed as mean ± standard deviation. As control, humanized mice not treated with pristane (NT) are included. N = 32 (14 CAR-Tregs, 8 UT-Tregs, 4 PBS, 6 NT). One-way ANOVA test with Tukey correction for multiple comparisons. B , C Percentage of total human CD45 + cells and human T and B cell sub-populations in the spleen and in the bone marrow in the different groups of treatment at sacrifice. T cells were defined as CD3 + cells. B cells were defined as: pre-B cells CD19 + CD20 - CD27 - cells, naïve B cells CD19 + CD20 + CD27 - cells, memory B cells CD19 + CD20 + CD27 + cells, plasmablasts CD19 + CD20 - CD27 + cells, plasma cells CD138 + cells. The results are expressed as mean ± standard deviation. As control, humanized mice not treated with pristane (NT) are included. N = 38 (16 CAR-Tregs, 11 UT-Tregs, 5 PBS, 6 NT). One-way ANOVA test with Tukey correction for multiple comparisons. Th exact p -values for each comparison are reported in the graphs.

Article Snippet: Fox19CAR engineered cells were identified using biotinylated recombinant human CD19 reagent (Miltenyi, 130-129-550, 1:67).

Techniques: Flow Cytometry, Standard Deviation, Control, Clinical Proteomics, Comparison

Journal: Nature Communications

Article Title: Regulatory T cells expressing CD19-targeted chimeric antigen receptor restore homeostasis in Systemic Lupus Erythematosus

doi: 10.1038/s41467-024-46448-9

Figure Lengend Snippet: A Schematic representation of the early treatment in a SGM-3 based humanized mouse model of SLE. B Longitudinal assessment of circulating Fox19CAR-Tregs and 19CAR-Tconvs after their injection 3 weeks after pristane (early treatment). N = 11 (5 CAR-Tregs, 6 CAR-Tconvs). C – E Absolute numbers of circulating human cells in the early treatment groups (3 weeks after pristane) after the injection of Fox19CAR-Tregs, 19CAR-Tconvs or PBS. Human cells were identified as total huCD45 + leukocytes, huCD45 + CD3 + T and huCD45 + CD19 + B lymphocytes, respectively. Absolute cell counts were assessed by flow cytometry. In B – E , black arrow indicates engineered cell injection. Green arrow denotes CAR-Treg second infusion. Results are expressed as mean ± standard deviation. A two-way ANOVA test with Tukey correction for multiple comparisons was employed. N = 17 (5 CAR-Tregs, 6 CAR-Tconvs, 6 PBS). * p -value 0.029. F Cytokine levels in peripheral blood of treated mice, injected 3 weeks after pristane. The amount of each cytokine was normalized across the various groups, scaled to range from 0 (minimum) to 1 (maximum). N = 17 (5 CAR-Tregs, 6 CAR-Tconvs, 4 PBS). G IL-10 levels in peripheral blood before and 7 days after treatment, 3 weeks after pristane. N = 14 (5 CAR-Tregs, 6 CAR-Tconvs, 4 PBS). Two-tailed Wilcoxon test for non-parametric paired data. * p -value 0.026, **** p -value < 0.0001.

Article Snippet: Fox19CAR engineered cells were identified using biotinylated recombinant human CD19 reagent (Miltenyi, 130-129-550, 1:67).

Techniques: Injection, Flow Cytometry, Standard Deviation, Two Tailed Test

Journal: Nature Communications

Article Title: Regulatory T cells expressing CD19-targeted chimeric antigen receptor restore homeostasis in Systemic Lupus Erythematosus

doi: 10.1038/s41467-024-46448-9

Figure Lengend Snippet: A Schematic representation of the late treatment in a SGM-3-based humanized mouse model of SLE. B Longitudinal assessment of circulating Fox19CAR-Tregs and 19CAR-Tconvs after their injection 8 weeks after pristane (late treatment). N = 10 (5 CAR-Tregs, 5 CAR-Tconvs). C Circulating human B lymphocytes (8 weeks after pristane) after the injection of Fox19CAR-Tregs, 19CAR-Tconvs or PBS. Human B lymphocytes were identified as huCD45 + CD19 + cells. Absolute cell counts were assessed by flow cytometry. N = 16 (5 CAR-Tregs, 5 CAR-Tconvs, 6 PBS). In B , C , black arrow indicates engineered cell injection. Results are expressed as mean ± standard deviation. Two-way ANOVA test with Tukey correction for multiple comparisons. D Cytokine levels in peripheral blood of treated mice. The amount of each cytokine was normalized, scaled to range from 0 (minimum) to 1 (maximum), and a color gradient was generated. For each group, the relative abundance of each cytokine is reported. N = 16 (5 late CAR-Tregs, 3 re-infused CAR-Tregs, 3 late CAR-Tconvs, 5 PBS). E IL-10 levels in peripheral blood before and 7 days after treatment 8 weeks after pristane. Results are expressed as mean ± standard deviation. N = 16 (5 late CAR-Tregs, 3 re-infused CAR-Tregs, 3 late CAR-Tconvs, 5 PBS). Two-way ANOVA with Tukey correction for multiple comparisons. * p -value 0.030, **** p -value < 0.0001.

Article Snippet: Fox19CAR engineered cells were identified using biotinylated recombinant human CD19 reagent (Miltenyi, 130-129-550, 1:67).

Techniques: Injection, Flow Cytometry, Standard Deviation, Generated

Journal: Molecular Cancer

Article Title: Safety and efficacy of a novel anti-CD19 chimeric antigen receptor T cell product targeting a membrane-proximal domain of CD19 with fast on- and off-rates against non-Hodgkin lymphoma: a first-in-human study

doi: 10.1186/s12943-023-01886-9

Figure Lengend Snippet: The h1218 antibody is specific for CD19 and recognizes a non-FMC63 membrane proximal epitope. A Schematic of FMC63 and h1218 antibodies binding sites on CD19. B Binding of h1218 to FMC63 bound-human CD19 complex. Sensor chips were coated with FMC63 Fc and CD19-ECD-Ck from 1700 to 2300 s. Additional FMC63 or h1218 antibody was added to the FMC63-bound sensor chip at 2300 s and monitored for further binding activities. C h1218 antibody binding test on wild-type HEK293T (CD19 negative), HEK293T cell expressing human CD19 (huCD19, HHH), and HEK293T cells expressing one of the three chimeric CD19 forms that had cynomolgus residues replacement at different region of CD19, respectively (CHH, HCH, and HHC). D Mutagenesis study to identify key residues corresponding to the h1218 CD19 epitope. E Quantification of IFNγ release on HEK293T cells expressing WT or mutant CD19 to identify additional key residues corresponding to the h1218 CD19 epitope. F Binding affinity of FMC63 and h1218 scFv to recombinant human CD19-ECD-Ck (M1-P278). Each line represents affinity measured at a different scFv concentration. All the experiments were repeated at least twice

Article Snippet: Lipid bilayers were first blocked with 5% casein for 20 min and then incubated with 6.3 nM streptavidin (ThermoFisher; Cat# 434301) for 15 min. After washing with imaging buffer (HEPES-buffered saline), lipids were incubated with biotinylated CD19 protein (R&D Systems; Cat#AVI9269) conjugated with Alexa Fluor 647 at room temperature for 30 min. After washing with the imaging buffer, lipid bilayers were blocked with 2.5 μM D-biotin to saturate the streptavidin-binding sites.

Techniques: Membrane, Binding Assay, Expressing, Mutagenesis, Recombinant, Concentration Assay

Journal: Molecular Cancer

Article Title: Safety and efficacy of a novel anti-CD19 chimeric antigen receptor T cell product targeting a membrane-proximal domain of CD19 with fast on- and off-rates against non-Hodgkin lymphoma: a first-in-human study

doi: 10.1186/s12943-023-01886-9

Figure Lengend Snippet: h1218-CART19 recognize and kill malignant B cells carrying FMC63-resistant CD19 mutations. A Schematic representation of B cell leukemia cells with point mutations in the membrane-distal CD19 domain (CD19 R163L or CD19 L174V ) as clinically identified post FMC63-CART19 treatment. B CD19 expression levels in Nalm6-CD19 KO, Nalm6 (wild type), Nalm6-CD19 L174V (left) and Nalm6-CD19 R163L (right) cell lines as measured by flow cytometry. C CART19 cytotoxicity against Nalm6-CD19 L174V and Nalm6-CD19 R163L at various effector to target (E:T) ratios ( n = 3 independent donors) (luciferase assay). D IL-2 and TNF cytokine release measured by ELISA 24 h after CART19 and cancer cell co-culture at an E:T ratio of 5:1 ( n = 2 donors). All values determined to be negative by comparison with the standard curve are shown as zero. E (Left) Schematic of the xenograft NSG mouse model: 1 × 10 6 luciferase + Nalm6-CD19 L174V cells and 0.75 × 10 6 UTD, FMC63-CART19, or h1218-CART19 cells were intravenous injected with a 5-day interval. (Right) Tumor burden of engrafted mice treated with UTD ( n = 5), FMC63-CART19 ( n = 5), or h1218-CART19 ( n = 5) as measured by bioluminescence imaging. The bold lines represent the median luminescence of each group. F Overall survival in each treatment group ( p = 0.0027). G Absolute cell counts of huCD45 + huCD3 + T cells in 100μL mouse blood on day 9. All bar graphs and cytotoxicity curves are presented as the mean ± SEM. Survival curves were compared using the log-rank (Mantel-Cox) test, and one-way ANOVA was performed with Tukey’s correction for multiple comparisons; **** p < 0.0001, *** p < 0.001, ** p < 0.01, and * p < 0.05. All bar graphs are presented as the mean ± SEM. All experiments were repeated at least twice

Article Snippet: Lipid bilayers were first blocked with 5% casein for 20 min and then incubated with 6.3 nM streptavidin (ThermoFisher; Cat# 434301) for 15 min. After washing with imaging buffer (HEPES-buffered saline), lipids were incubated with biotinylated CD19 protein (R&D Systems; Cat#AVI9269) conjugated with Alexa Fluor 647 at room temperature for 30 min. After washing with the imaging buffer, lipid bilayers were blocked with 2.5 μM D-biotin to saturate the streptavidin-binding sites.

Techniques: Membrane, Expressing, Flow Cytometry, Luciferase, Enzyme-linked Immunosorbent Assay, Co-Culture Assay, Comparison, Injection, Imaging

Journal: Molecular Cancer

Article Title: Safety and efficacy of a novel anti-CD19 chimeric antigen receptor T cell product targeting a membrane-proximal domain of CD19 with fast on- and off-rates against non-Hodgkin lymphoma: a first-in-human study

doi: 10.1186/s12943-023-01886-9

Figure Lengend Snippet: h1218-CART19 demonstrates enhanced efficacy than FMC63-CART19. A Schematic of h1218-CART19 targeting the membrane-proximal domain of CD19 in B cell lymphoma/leukemia as compared to standard FMC63-CART19. B Nalm6 tumor killing by UTD, h1218-CART19, or FMC63-CART19 cells at various E:T ratios ( n = 3 donors) by flow cytometry. C IFNγ release as measured by ELISA upon stimulation of CD19-expressing tumor cells (Raji, Pfeiffer, Toledo, and Nalm6) at an E:T ratio of 3:1 ( n = 2 donors). D (Left) Schematic of the in vivo experiment: either 1.5 × 10 6 UTD or CART19 cells were infused 7 days after intravenous injection of luciferase + Raji or lNalm6 cells. Tumor progression in mice bearing Raji (middle) or Nalm6 (right) cells is shown. E Quantification of Nalm6 fold change over 14 days in the presence of UTD, FMC63-CART19, or h1218-CART19 at low-E:T ratio model ( n = 2 donors) using flow cytometry. F (Left) Schematic of the in vivo xenograft model: 0.75 × 10 6 UTD or CAR T cells were infused intravenously 5 days after luciferase + Nalm6 engraftment. (Middle) Tumor progression over time in mice bearing Nalm6 cells treated with UTD ( n = 3), FMC63-CART19 ( n = 7), or h1218-CART19 ( n = 7) measured by luminescence. Bolded lines represent the median tumor burden in the corresponding group. (Right) Tumor burden on day 42 after CART19 injection. G Overall survival in mice bearing Nalm6. H (Left) CAR T cell expansion kinetics in the peripheral blood after CART19 injection. Bolded lines represent the median CAR T expansion. (Right) Quantification of CAR T cells in the blood 14 days after CART19 injection using flow cytometry. All bar graphs and cytotoxicity curves are presented as the mean ± SEM. Student's t-test was used to compare two groups; one-way ANOVA was performed with Tukey’s correction for multiple comparisons; survival curves were compared using the log-rank (Mantel-Cox) test; **** p < 0.0001, *** p < 0.001, ** p < 0.01, and * p < 0.05. All bar graphs are presented as the mean ± SEM. All experiments were repeated at least twice

Article Snippet: Lipid bilayers were first blocked with 5% casein for 20 min and then incubated with 6.3 nM streptavidin (ThermoFisher; Cat# 434301) for 15 min. After washing with imaging buffer (HEPES-buffered saline), lipids were incubated with biotinylated CD19 protein (R&D Systems; Cat#AVI9269) conjugated with Alexa Fluor 647 at room temperature for 30 min. After washing with the imaging buffer, lipid bilayers were blocked with 2.5 μM D-biotin to saturate the streptavidin-binding sites.

Techniques: Membrane, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Expressing, In Vivo, Injection, Luciferase

Journal: Molecular Cancer

Article Title: Safety and efficacy of a novel anti-CD19 chimeric antigen receptor T cell product targeting a membrane-proximal domain of CD19 with fast on- and off-rates against non-Hodgkin lymphoma: a first-in-human study

doi: 10.1186/s12943-023-01886-9

Figure Lengend Snippet: h1218-CART19 demonstrates lower avidity and less activation-induced cell death than FMC63-CART19. A Quantification of UTD, FMC63-CART19, and h1218-CART19 binding avidity to Nalm6 after 15-min co-culture ( n = 2 donors) by Lumicks analysis. B Representative confocal microscopy images of F-actin, CD19, perforin, and phosphorylated-CD3ζ (pCD3ζ) (CAR) expressed in FMC63-CART19 or h1218-CART19 cells when engaged with biotinylated CD19 protein. C Quantification of F-actin, perforin polarization, and pCD3ζ in FMC63-CART19 or h1219-CART19 cells engaged with biotinylated CD19 protein ( n = 2 donors). In total, 200 events were recorded for each group. D (Left) Representative flow cytometric analysis of Caspase3/7 + FMC63-CART19 and h1218-CART19 cells with and without 4-h stimulation by Nalm6. Caspase3/7 + population of CART19 cells is boxed in red. (Middle) Baseline level of Caspase3/7 + population in FMC63- or h1218-CART19 cells at 0 h. (Right) Differential increase in the Caspase3/7 + population after 4-h stimulation. All graphs are represented as the mean ± SEM. Student's t-test was used to compare two groups; one-way ANOVA was performed with Tukey’s correction for multiple comparisons; **** p < 0.0001, *** p < 0.001, ** p < 0.01, and * p < 0.05. All bar graphs are presented as the mean ± SEM. All experiments were repeated at least twice

Article Snippet: Lipid bilayers were first blocked with 5% casein for 20 min and then incubated with 6.3 nM streptavidin (ThermoFisher; Cat# 434301) for 15 min. After washing with imaging buffer (HEPES-buffered saline), lipids were incubated with biotinylated CD19 protein (R&D Systems; Cat#AVI9269) conjugated with Alexa Fluor 647 at room temperature for 30 min. After washing with the imaging buffer, lipid bilayers were blocked with 2.5 μM D-biotin to saturate the streptavidin-binding sites.

Techniques: Activation Assay, Binding Assay, Co-Culture Assay, Confocal Microscopy