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Cell Signaling Technology Inc anti egfr monoclonal rat antibody

CD44 and <t>EGFR</t> overexpression in VA-ES-BJ cells and in human ES. (a) The cell surface expression of CD44 and EGFR on VA-ES-BJ cells measured by flow cytometry. The corresponding isotype antibodies were used as controls. (b) Representative images from three VA-ES-BJ xenograft tumor sections after H&E staining (left), immunostaining for CD44 (middle) or EGFR (right) at 20× magnification. Scale bar = 100 μm. Representative images from human ES tumor tissue sections after H&E staining (c), immunostaining for CD44 (d) and EGFR (e) at 20× magnification. Human ES tumor tissue samples were resected from four ES patients. Scale bar = 100 μm.

Anti Egfr Monoclonal Rat Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 91/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/cd44+rat+monoclonal+antibody/CD44+Rat+mAb/pmc11063645-71-14-20
Average 91 stars, based on 2 article reviews

anti egfr monoclonal rat antibody - by Bioz Stars, 2026-09

91/100 stars

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1) Product Images from "Photoimmunotheranostics of epithelioid sarcoma by targeting CD44 or EGFR"

Article Title: Photoimmunotheranostics of epithelioid sarcoma by targeting CD44 or EGFR

Journal: Translational Oncology

doi: 10.1016/j.tranon.2024.101966

CD44 and EGFR overexpression in VA-ES-BJ cells and in human ES. (a) The cell surface expression of CD44 and EGFR on VA-ES-BJ cells measured by flow cytometry. The corresponding isotype antibodies were used as controls. (b) Representative images from three VA-ES-BJ xenograft tumor sections after H&E staining (left), immunostaining for CD44 (middle) or EGFR (right) at 20× magnification. Scale bar = 100 μm. Representative images from human ES tumor tissue sections after H&E staining (c), immunostaining for CD44 (d) and EGFR (e) at 20× magnification. Human ES tumor tissue samples were resected from four ES patients. Scale bar = 100 μm.

Figure Legend Snippet: CD44 and EGFR overexpression in VA-ES-BJ cells and in human ES. (a) The cell surface expression of CD44 and EGFR on VA-ES-BJ cells measured by flow cytometry. The corresponding isotype antibodies were used as controls. (b) Representative images from three VA-ES-BJ xenograft tumor sections after H&E staining (left), immunostaining for CD44 (middle) or EGFR (right) at 20× magnification. Scale bar = 100 μm. Representative images from human ES tumor tissue sections after H&E staining (c), immunostaining for CD44 (d) and EGFR (e) at 20× magnification. Human ES tumor tissue samples were resected from four ES patients. Scale bar = 100 μm.

Techniques Used: Over Expression, Expressing, Flow Cytometry, Staining, Immunostaining

Concentration and exposure-dependence and target-specific cell death of CD44-IR700 and EGFR-IR700. CD44-IR700-mediated phototoxicity was dependent on the concentration of CD44-IR700 (a) or exposure dose (b). (c) CD44-specific cell death only occurred when VA-ES-BJ cells were exposed to CD44-IR700 and light irradiation. EGFR-IR700-mediated phototoxicity was dependent on the concentration of EGFR-IR700 (d) or exposure dose (e). (f) EGFR-specific cell death only occurred when VA-ES-BJ cells were exposed to both EGFR-IR700 and light irradiation.

Figure Legend Snippet: Concentration and exposure-dependence and target-specific cell death of CD44-IR700 and EGFR-IR700. CD44-IR700-mediated phototoxicity was dependent on the concentration of CD44-IR700 (a) or exposure dose (b). (c) CD44-specific cell death only occurred when VA-ES-BJ cells were exposed to CD44-IR700 and light irradiation. EGFR-IR700-mediated phototoxicity was dependent on the concentration of EGFR-IR700 (d) or exposure dose (e). (f) EGFR-specific cell death only occurred when VA-ES-BJ cells were exposed to both EGFR-IR700 and light irradiation.

Techniques Used: Concentration Assay, Irradiation

Preferential accumulation of CD44-IR700 or EGFR-IR700 in VA-ES-BJ tumors. NIR fluorescence imaging of nude mice bearing bilateral VA-ES-BJ tumors over a 24-h period. 100 μg of CD44-IR700 (a) or EGFR-IR700 (b) or IgG-IR700 (c) antibody conjugate was injected i.v. (d) Plots of tumor to normal (T/N) ratios measured at 0 min, 1 h, 6 h and 24 h p.i. ROIs of tumors and normal tissue were drawn on in vivo NIR fluorescence images. T/N ratios were derived from the MFI of ROIs and represented as Mean ± SD for all three groups ( n = 10 per group). Comparisons of T/N ratios between the groups at 6 h p.i. and 24 h p.i. are shown on the right, ns: no significance, * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. (e) Ex vivo fluorescence images of whole tumors resected at 24 h p.i. (f) The MFI of resected tumors at 24 h p.i. in different groups. Values represent Mean ± SD from six tumors per group ( n = 6). (g) Bio-distribution of antibody conjugates in main organs and tumors resected at 24 h p.i. H, heart; Li, liver; Sp, spleen; Lu, lung; K, kidney; In, intestine; St, stomach; Mu, muscle; Bn, bone; Bl, blood; T, tumor. Values (Mean ± SD) are normalized to % injected dose/g (% ID /g) from three mice per group ( n = 3). Only comparisons with p value less than 0.05 displayed, * P < 0.05, *** P < 0.001, **** P < 0.0001.

Figure Legend Snippet: Preferential accumulation of CD44-IR700 or EGFR-IR700 in VA-ES-BJ tumors. NIR fluorescence imaging of nude mice bearing bilateral VA-ES-BJ tumors over a 24-h period. 100 μg of CD44-IR700 (a) or EGFR-IR700 (b) or IgG-IR700 (c) antibody conjugate was injected i.v. (d) Plots of tumor to normal (T/N) ratios measured at 0 min, 1 h, 6 h and 24 h p.i. ROIs of tumors and normal tissue were drawn on in vivo NIR fluorescence images. T/N ratios were derived from the MFI of ROIs and represented as Mean ± SD for all three groups ( n = 10 per group). Comparisons of T/N ratios between the groups at 6 h p.i. and 24 h p.i. are shown on the right, ns: no significance, * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. (e) Ex vivo fluorescence images of whole tumors resected at 24 h p.i. (f) The MFI of resected tumors at 24 h p.i. in different groups. Values represent Mean ± SD from six tumors per group ( n = 6). (g) Bio-distribution of antibody conjugates in main organs and tumors resected at 24 h p.i. H, heart; Li, liver; Sp, spleen; Lu, lung; K, kidney; In, intestine; St, stomach; Mu, muscle; Bn, bone; Bl, blood; T, tumor. Values (Mean ± SD) are normalized to % injected dose/g (% ID /g) from three mice per group ( n = 3). Only comparisons with p value less than 0.05 displayed, * P < 0.05, *** P < 0.001, **** P < 0.0001.

Techniques Used: Fluorescence, Imaging, Injection, In Vivo, Derivative Assay, Ex Vivo

In vivo CD44-IR700 or EGFR-IR700-PIT. (a) Growth curve of VA-ES-BJ tumors. 100 μg of CD44-IR700 or EGFR-IR700 or IgG-IR700 was injected i.v. on day 0 and on day 7. All the groups were monitored for three weeks after injection except for two weeks in PBS group. PIT groups received light exposure at 200 J/cm 2 , 24 h after each injection (pointed by blue arrows) and mice in the no PIT groups were shielded from light. Values represent Mean ± SEM from at least four mice per group ( n ≥ 4). (b) The comparisons of tumor volumes between the groups at each monitoring point, ns: no significance, * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. (c) Photographs of representative VA-ES-BJ tumors harvested at the end of treatment.

Figure Legend Snippet: In vivo CD44-IR700 or EGFR-IR700-PIT. (a) Growth curve of VA-ES-BJ tumors. 100 μg of CD44-IR700 or EGFR-IR700 or IgG-IR700 was injected i.v. on day 0 and on day 7. All the groups were monitored for three weeks after injection except for two weeks in PBS group. PIT groups received light exposure at 200 J/cm 2 , 24 h after each injection (pointed by blue arrows) and mice in the no PIT groups were shielded from light. Values represent Mean ± SEM from at least four mice per group ( n ≥ 4). (b) The comparisons of tumor volumes between the groups at each monitoring point, ns: no significance, * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. (c) Photographs of representative VA-ES-BJ tumors harvested at the end of treatment.

Techniques Used: In Vivo, Injection

Short-term effects of PIT on CD44 and EGFR. (a) Immunoblots probing CD44 and EGFR protein in VA-ES-BJ tumors. (b) Representative flow cytometry profiles of CD44 and EGFR expression in tumors after receiving PITs. 100 μg of CD44-IR700 or EGFR-IR700 or 100 μl of PBS was injected i.v. PIT groups received light exposure at 200 J/cm 2 24 h after injection and mice in the PBS group were shielded from light. VA-ES-BJ tumors were harvested three days after single PIT. Tumors were dissociated into a single cell suspension and only live cells were included in the flow cytometry analysis.

Figure Legend Snippet: Short-term effects of PIT on CD44 and EGFR. (a) Immunoblots probing CD44 and EGFR protein in VA-ES-BJ tumors. (b) Representative flow cytometry profiles of CD44 and EGFR expression in tumors after receiving PITs. 100 μg of CD44-IR700 or EGFR-IR700 or 100 μl of PBS was injected i.v. PIT groups received light exposure at 200 J/cm 2 24 h after injection and mice in the PBS group were shielded from light. VA-ES-BJ tumors were harvested three days after single PIT. Tumors were dissociated into a single cell suspension and only live cells were included in the flow cytometry analysis.

Techniques Used: Western Blot, Flow Cytometry, Expressing, Injection, Suspension

Flow Cytometry:

Article Title: lnc-PKD2-2-3, identified by long non-coding RNA expression profiling, is associated with pejorative tumor features and poor prognosis, enhances cancer stemness and may serve as cancer stem-cell marker in cholangiocarcinoma
Article Snippet: In addition, CD44 + CD133 + cell proportions at 72 h were detected by flow cytometry using a FACSCalibur (BD Biosciences) and analyzed using Flowjo Software 7.6 (FlowJo, LLC). .. The antibodies used in flow cytometry were CD133 mouse monoclonal antibody (cat. no. 38725, flow-specific; Alexa Fluor ® 488-conjugated; Cell Signaling Technology, Inc.) and CD44 rat monoclonal antibody (cat. no. 80813; allophycocyanin-conjugated; Cell Signaling Technology, Inc.). ..

Image Search Results

Estimated copies per cell of PIM1 and PIM2 protein from quantitative proteomics analysis of ( A ) OT1 CD8 T cells stimulated with SIINFEKL peptide for indicated times from published dataset or ( B ) naive ex vivo and 24 hr αCD3/αCD28 (TCR) activated WT CD8 T cells (see ( G, H )) for further details. ( C ) Fragments per kilobase million (FPKM) of Pim1 , Pim2, and Pim3 mRNA from published bulk RNAseq analysis of naive and 24 hr gp33-41 peptide stimulated P14 CD8 T cells. Lymph node cell suspensions from C57BL/6 (WT) and Pim1 KO/ Pim2 KO (Pim dKO) mice were activated for 24 hr with αCD3/αCD28 (both 0.5 µg/mL) and CD4 and CD8 T cell ( D ) FSC-A SSC-A profiles, ( E ) expression of surface activation markers (CD25, CD44, CD71) or CD8 T cell intracellular IFNγ were measured by flow cytometry. ( F ) Lymph node single-cell suspensions from WT and Pim dKO mice were labelled with CellTrace Violet (CTV), activated with αCD3/αCD28 (both 0.5 µg/mL) and CD4 and CD8 T cell CTV proliferation profiles were measured at indicated time points. ( G, H ) Lymph node cell suspensions from WT and Pim dKO mice were stimulated for 24 hr with αCD3/αCD28 (both 0.5 µg/mL) and activated CD4 and CD8 T cells were sorted for analysis by quantitative proteomics. Data was analysed using proteomic ruler method to estimate protein copy number per cell. An interactive version of the proteomics expression data is available for exploration on the Immunological Proteome Resource website: immpres.co.uk ( G ) Total protein content (µg/million cells) (one-way ANOVA), ( H ) Volcano plots of p-value (-log 10 ) versus fold-change (log 2 ) in protein copy number between Pim dKO and WT. Horizontal dotted line represents multi-test correction cut-off of q=0.05, vertical dotted line shows 1.5-fold change. Phosphoribosyl Pyrophosphate synthase 1 like 1 (Prps1l1), was found to be higher in Pim dKO CD8 T cells, but was a low confidence quantification (based on only two unique peptides) with no known function in T cells. Lymph node single-cell suspensions from WT and Pim dKO mice were labelled with CellTrace Violet (CTV) and ( I ) cells were cultured in IL-7 (5 ng/mL) +/- rapamycin (20 nM) and CD8 T cell numbers measured over time or ( J ) cells were activated with αCD3/αCD28 (both 0.5 µg/mL) +/- rapamycin (20 nM) and CD8 T cell mean division number was calculated over time (two-way ANOVA). Symbols in bar charts represent biological replicates, symbols in ( I ) represent the mean. Error bars show mean ± S.D. Flow cytometry dot plots and histograms are representative of ( D, E ) n=3, except for IFNγ staining which is n=2, ( F ) n=5, or show pooled data from ( I ) n=3–4 and ( J ) n=5 biological replicates, with data collected over at least two independent experiments. Quantitative proteomics was performed on biological triplicates. Figure 1—source data 1. Raw values plotted in .

Journal: eLife

Article Title: PIM kinase control of CD8 T cell protein synthesis and cell trafficking

doi: 10.7554/eLife.98622

Figure Lengend Snippet: Estimated copies per cell of PIM1 and PIM2 protein from quantitative proteomics analysis of ( A ) OT1 CD8 T cells stimulated with SIINFEKL peptide for indicated times from published dataset or ( B ) naive ex vivo and 24 hr αCD3/αCD28 (TCR) activated WT CD8 T cells (see ( G, H )) for further details. ( C ) Fragments per kilobase million (FPKM) of Pim1 , Pim2, and Pim3 mRNA from published bulk RNAseq analysis of naive and 24 hr gp33-41 peptide stimulated P14 CD8 T cells. Lymph node cell suspensions from C57BL/6 (WT) and Pim1 KO/ Pim2 KO (Pim dKO) mice were activated for 24 hr with αCD3/αCD28 (both 0.5 µg/mL) and CD4 and CD8 T cell ( D ) FSC-A SSC-A profiles, ( E ) expression of surface activation markers (CD25, CD44, CD71) or CD8 T cell intracellular IFNγ were measured by flow cytometry. ( F ) Lymph node single-cell suspensions from WT and Pim dKO mice were labelled with CellTrace Violet (CTV), activated with αCD3/αCD28 (both 0.5 µg/mL) and CD4 and CD8 T cell CTV proliferation profiles were measured at indicated time points. ( G, H ) Lymph node cell suspensions from WT and Pim dKO mice were stimulated for 24 hr with αCD3/αCD28 (both 0.5 µg/mL) and activated CD4 and CD8 T cells were sorted for analysis by quantitative proteomics. Data was analysed using proteomic ruler method to estimate protein copy number per cell. An interactive version of the proteomics expression data is available for exploration on the Immunological Proteome Resource website: immpres.co.uk ( G ) Total protein content (µg/million cells) (one-way ANOVA), ( H ) Volcano plots of p-value (-log 10 ) versus fold-change (log 2 ) in protein copy number between Pim dKO and WT. Horizontal dotted line represents multi-test correction cut-off of q=0.05, vertical dotted line shows 1.5-fold change. Phosphoribosyl Pyrophosphate synthase 1 like 1 (Prps1l1), was found to be higher in Pim dKO CD8 T cells, but was a low confidence quantification (based on only two unique peptides) with no known function in T cells. Lymph node single-cell suspensions from WT and Pim dKO mice were labelled with CellTrace Violet (CTV) and ( I ) cells were cultured in IL-7 (5 ng/mL) +/- rapamycin (20 nM) and CD8 T cell numbers measured over time or ( J ) cells were activated with αCD3/αCD28 (both 0.5 µg/mL) +/- rapamycin (20 nM) and CD8 T cell mean division number was calculated over time (two-way ANOVA). Symbols in bar charts represent biological replicates, symbols in ( I ) represent the mean. Error bars show mean ± S.D. Flow cytometry dot plots and histograms are representative of ( D, E ) n=3, except for IFNγ staining which is n=2, ( F ) n=5, or show pooled data from ( I ) n=3–4 and ( J ) n=5 biological replicates, with data collected over at least two independent experiments. Quantitative proteomics was performed on biological triplicates. Figure 1—source data 1. Raw values plotted in .

Article Snippet: Antibody , anti-mouse CD44 (Rat, monoclonal, IM7) , Thermo Fisher Scientific/eBioscience , Cat # 47-0441-82, RRID: AB_1272244 , cell surface stain 1:200, APC eF780.

Techniques: Quantitative Proteomics, Ex Vivo, Expressing, Activation Assay, Flow Cytometry, Cell Culture, Staining

( A ) Estimated copies per cell of PIM1 and PIM2 protein from published quantitative proteomics analysis ; of CD8 T cells expanded in IL-2 or IL-15 as outlined in . ( B–D, F, G ) WT (Ly5.1) and Pim dKO lymph node or spleen single-cell suspensions were mixed at a 50:50 ratio of T cells, activated for 2 days with αCD3/αCD28 (both 0.5 µg/mL) and IL-2 (20 ng/mL), washed then split into fresh medium containing IL-2 (20 ng/mL) daily (as per ). Some of the mixed cell suspensions were also cultured in IL-7 (5 ng/mL) to sustain a naive T cell reference. ( B ) WT and Pim dKO CTL were treated 1 hr +/- Jak1/3 inhibitor Tofacitinib (100 nM; negative control) before pSTAT5 Y694 expression was measured on day 3 and 6 of culture, ( C ) surface CD25 expression was measured on days 3 and 6 of culture, ( D ) CD8 T cell number vs time was calculated, ( F ) CD8 T cell FSC-A, SSC-A and surface activation markers (CD44, CD71) were measured on days 3 and 6 of culture ( G ) expression of adhesion molecule CD62L was measured daily. ( E ) WT and Pim dKO T cells were activated and expanded with IL-2 as per ( B-D ) and ( F, G ) except in separate cultures and % live cells (PI-ve) was assessed on days 4 and 6 (two-way ANOVA). Symbols in bar charts represent biological replicates, symbols in ( D ) represent the mean. Error bars show mean ± S.D. Data are representative of ( B, G ) n=4, ( C, F ) n=6 or show pooled data from ( D ) n=4, ( E ) n=6 biological replicates with data collected over at least two independent experiments. Figure 3—source data 1. Raw values plotted in .

Journal: eLife

Article Title: PIM kinase control of CD8 T cell protein synthesis and cell trafficking

doi: 10.7554/eLife.98622

Figure Lengend Snippet: ( A ) Estimated copies per cell of PIM1 and PIM2 protein from published quantitative proteomics analysis ; of CD8 T cells expanded in IL-2 or IL-15 as outlined in . ( B–D, F, G ) WT (Ly5.1) and Pim dKO lymph node or spleen single-cell suspensions were mixed at a 50:50 ratio of T cells, activated for 2 days with αCD3/αCD28 (both 0.5 µg/mL) and IL-2 (20 ng/mL), washed then split into fresh medium containing IL-2 (20 ng/mL) daily (as per ). Some of the mixed cell suspensions were also cultured in IL-7 (5 ng/mL) to sustain a naive T cell reference. ( B ) WT and Pim dKO CTL were treated 1 hr +/- Jak1/3 inhibitor Tofacitinib (100 nM; negative control) before pSTAT5 Y694 expression was measured on day 3 and 6 of culture, ( C ) surface CD25 expression was measured on days 3 and 6 of culture, ( D ) CD8 T cell number vs time was calculated, ( F ) CD8 T cell FSC-A, SSC-A and surface activation markers (CD44, CD71) were measured on days 3 and 6 of culture ( G ) expression of adhesion molecule CD62L was measured daily. ( E ) WT and Pim dKO T cells were activated and expanded with IL-2 as per ( B-D ) and ( F, G ) except in separate cultures and % live cells (PI-ve) was assessed on days 4 and 6 (two-way ANOVA). Symbols in bar charts represent biological replicates, symbols in ( D ) represent the mean. Error bars show mean ± S.D. Data are representative of ( B, G ) n=4, ( C, F ) n=6 or show pooled data from ( D ) n=4, ( E ) n=6 biological replicates with data collected over at least two independent experiments. Figure 3—source data 1. Raw values plotted in .

Article Snippet: Antibody , anti-mouse CD44 (Rat, monoclonal, IM7) , Thermo Fisher Scientific/eBioscience , Cat # 47-0441-82, RRID: AB_1272244 , cell surface stain 1:200, APC eF780.

Techniques: Quantitative Proteomics, Cell Culture, Negative Control, Expressing, Activation Assay

In vivo effect of IMT504 on Wnt1 or GLAST-traced bone marrow mononuclear cells. ( A ) Statistical comparisons of numbers of CFU-Fs, Tom + or Tom − , obtained from BM-MNCs of Wnt1 Cre ; Rosa26 Tom which were or not treated with TAA during 2 weeks, and were s.c. injected or not with IMT504. ( B ) Statistical comparisons of percentage of CFU-Fs Tom + . ( A, B ) Dunn`s multiple comparison test; n = 6. ( C ) Microphotographs showing representative colonies obtained from naïve, TAA 2w and TAA 2w + IMT504 treated mice. Scale bar: 50 μm. ( D , left) Statistical comparisons of percentage of CD44 + Tom + cells within the bone marrow after 2 weeks of TAA and/or IMT504 treatments, as measured by flow cytometry. ( D , right) Statistical comparisons of percentage of Ki67 + cells among the total of CD44 + Tom + cells within the bone marrow from GLAST CreERT2 ; Rosa26 Tom (Tx P2) after 2 weeks of TAA and IMT504/saline treatments, as measured by flow cytometry. ( E ) Statistical comparisons of percentage of CD44 + Tom + cells within the peripheral blood after 2 weeks of TAA and IMT504/saline treatments, as measured by flow cytometry. ( D, E ) Turkey`s multiple comparison test; n = 4. ( A, B,D, E ) * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001

Journal: Stem Cell Research & Therapy

Article Title: The antifibrotic potential of IMT504: modulation of GLAST + Wnt1 + bone marrow stromal progenitors and hepatic microenvironment

doi: 10.1186/s13287-024-03896-w

Figure Lengend Snippet: In vivo effect of IMT504 on Wnt1 or GLAST-traced bone marrow mononuclear cells. ( A ) Statistical comparisons of numbers of CFU-Fs, Tom + or Tom − , obtained from BM-MNCs of Wnt1 Cre ; Rosa26 Tom which were or not treated with TAA during 2 weeks, and were s.c. injected or not with IMT504. ( B ) Statistical comparisons of percentage of CFU-Fs Tom + . ( A, B ) Dunn`s multiple comparison test; n = 6. ( C ) Microphotographs showing representative colonies obtained from naïve, TAA 2w and TAA 2w + IMT504 treated mice. Scale bar: 50 μm. ( D , left) Statistical comparisons of percentage of CD44 + Tom + cells within the bone marrow after 2 weeks of TAA and/or IMT504 treatments, as measured by flow cytometry. ( D , right) Statistical comparisons of percentage of Ki67 + cells among the total of CD44 + Tom + cells within the bone marrow from GLAST CreERT2 ; Rosa26 Tom (Tx P2) after 2 weeks of TAA and IMT504/saline treatments, as measured by flow cytometry. ( E ) Statistical comparisons of percentage of CD44 + Tom + cells within the peripheral blood after 2 weeks of TAA and IMT504/saline treatments, as measured by flow cytometry. ( D, E ) Turkey`s multiple comparison test; n = 4. ( A, B,D, E ) * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001

Article Snippet: Other markers used in flow cytometry experiments were: mouse monoclonal anti-Albumin (Santa Cruz Biotechnology; sc-374670), rat monoclonal anti-CD31 (BD Pharmingen; 553370), rat monoclonal anti-CD44-APC antibody (BD Pharmingen; 559250) rabbit polyclonal anti-CD133 (Abcam; ab19898), goat anti-Ep-CAM (Santa Cruz Biotechnology; sc-23788).

Techniques: In Vivo, Injection, Comparison, Flow Cytometry, Saline

Canonical TRα signaling limits the induction and shapes the phenotype of regulatory T cells (A) Percentage of Foxp3+ and (B) CTLA4+CD4 + T cells was measured in spleens of naive mice by flow cytometry. (C) IL-10 was determined in serum of naive mice via Luminex. (D) Naive CD25 − CD4 + T cells were differentiated into Treg by in vitro polarization assay. Exemplary dot plots and frequency of Foxp3+CD25 + induced Treg at day 3 are depicted. (E) Frequency of Treg was examined in the thymus of naive TRαGS and TRαWT mice. (F) CD25 + Treg ( n = 4) of TRαGS and TRαWT mice were isolated from spleen. Exemplary dot plots show frequency of Foxp3+ cells of CD25 + CD4 + T cells in spleens of naive TRαWT and TRαGS mice. (G) Transcriptome of isolated Treg was analyzed by RNA-sequencing and principal-component analysis of differently expressed genes was performed assessing changes between genotypes. (H) Heatmap shows 20 most down and (I) up regulated genes in TRαGS Treg compared to TRαWT cells. (J) Differently expressed genes were used for overrepresentation analysis and 10 most enriched GO-terms are shown. (K) Frequency of CD62 low CD44 + effector Foxp3+ Treg and (L) CD62 low CD44 + effector Foxp3- CD4 + T cells was determined in spleen of naive mice by flow cytometry. (M) Markers of Treg function were determined on splenic Foxp3+ Treg in naive mice by flow cytometry. (N) Proliferation of splenic CD25 + Treg following in vitro expansion for 72 h was examined using CFSE cell tracer. (O) Suppressive activity of TRαGS and TRαWT Treg was determined in an in vitro suppression assay using indicated ratios of T responder cells (Tresp; TRαWT CD4 + T cells) and CD25 + Treg. (P) Percentage of CD25 + Foxp3+ Treg was analyzed by flow cytometry in spleen of naive mice. (Q) Differential expression of genes involved in T cell migration from RNA-sequencing analysis of isolated TRαWT and TRαGS Treg. Data are shown as mean ± SD pooled from 1 to 7 independent experiments containing at least 3 biological replicates. For statistical analysis data were tested for normal distribution using D’Agostino and Pearson omnibus normality test. Means were compared by unpaired t test or Mann-Whitney U test, comparison of multiple groups was done by two-way ANOVA and post hoc Tukey test. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001.

Journal: iScience

Article Title: Lack of canonical thyroid hormone receptor α signaling changes regulatory T cell phenotype in female mice

doi: 10.1016/j.isci.2024.110547

Figure Lengend Snippet: Canonical TRα signaling limits the induction and shapes the phenotype of regulatory T cells (A) Percentage of Foxp3+ and (B) CTLA4+CD4 + T cells was measured in spleens of naive mice by flow cytometry. (C) IL-10 was determined in serum of naive mice via Luminex. (D) Naive CD25 − CD4 + T cells were differentiated into Treg by in vitro polarization assay. Exemplary dot plots and frequency of Foxp3+CD25 + induced Treg at day 3 are depicted. (E) Frequency of Treg was examined in the thymus of naive TRαGS and TRαWT mice. (F) CD25 + Treg ( n = 4) of TRαGS and TRαWT mice were isolated from spleen. Exemplary dot plots show frequency of Foxp3+ cells of CD25 + CD4 + T cells in spleens of naive TRαWT and TRαGS mice. (G) Transcriptome of isolated Treg was analyzed by RNA-sequencing and principal-component analysis of differently expressed genes was performed assessing changes between genotypes. (H) Heatmap shows 20 most down and (I) up regulated genes in TRαGS Treg compared to TRαWT cells. (J) Differently expressed genes were used for overrepresentation analysis and 10 most enriched GO-terms are shown. (K) Frequency of CD62 low CD44 + effector Foxp3+ Treg and (L) CD62 low CD44 + effector Foxp3- CD4 + T cells was determined in spleen of naive mice by flow cytometry. (M) Markers of Treg function were determined on splenic Foxp3+ Treg in naive mice by flow cytometry. (N) Proliferation of splenic CD25 + Treg following in vitro expansion for 72 h was examined using CFSE cell tracer. (O) Suppressive activity of TRαGS and TRαWT Treg was determined in an in vitro suppression assay using indicated ratios of T responder cells (Tresp; TRαWT CD4 + T cells) and CD25 + Treg. (P) Percentage of CD25 + Foxp3+ Treg was analyzed by flow cytometry in spleen of naive mice. (Q) Differential expression of genes involved in T cell migration from RNA-sequencing analysis of isolated TRαWT and TRαGS Treg. Data are shown as mean ± SD pooled from 1 to 7 independent experiments containing at least 3 biological replicates. For statistical analysis data were tested for normal distribution using D’Agostino and Pearson omnibus normality test. Means were compared by unpaired t test or Mann-Whitney U test, comparison of multiple groups was done by two-way ANOVA and post hoc Tukey test. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001.

Article Snippet: Rat anti-CD44 monoclonal antibody; FITC , BD Biosciences , Cat# 561859; RRID: AB_10894581.

Techniques: Flow Cytometry, Luminex, In Vitro, Isolation, RNA Sequencing Assay, Activity Assay, Suppression Assay, Expressing, Migration, MANN-WHITNEY, Comparison

Journal: iScience

Article Title: Lack of canonical thyroid hormone receptor α signaling changes regulatory T cell phenotype in female mice

doi: 10.1016/j.isci.2024.110547

Figure Lengend Snippet:

Article Snippet: Rat anti-CD44 monoclonal antibody; FITC , BD Biosciences , Cat# 561859; RRID: AB_10894581.

Techniques: Functional Assay, Recombinant, Cell Isolation, Staining, Luciferase, Plasmid Preparation, Software, Flow Cytometry, Electroporation

Journal: Translational Oncology

Article Title: Photoimmunotheranostics of epithelioid sarcoma by targeting CD44 or EGFR

doi: 10.1016/j.tranon.2024.101966

Figure Lengend Snippet: CD44 and EGFR overexpression in VA-ES-BJ cells and in human ES. (a) The cell surface expression of CD44 and EGFR on VA-ES-BJ cells measured by flow cytometry. The corresponding isotype antibodies were used as controls. (b) Representative images from three VA-ES-BJ xenograft tumor sections after H&E staining (left), immunostaining for CD44 (middle) or EGFR (right) at 20× magnification. Scale bar = 100 μm. Representative images from human ES tumor tissue sections after H&E staining (c), immunostaining for CD44 (d) and EGFR (e) at 20× magnification. Human ES tumor tissue samples were resected from four ES patients. Scale bar = 100 μm.

Article Snippet: An anti-CD44 monoclonal mouse antibody (clone 156-3C11, Cell Signaling, Danvers, MA, USA) and an anti-EGFR monoclonal rat antibody (clone D38B1, Cell Signaling) were used for CD44 and EGFR immunostaining, respectively.

Techniques: Over Expression, Expressing, Flow Cytometry, Staining, Immunostaining

Journal: Translational Oncology

Article Title: Photoimmunotheranostics of epithelioid sarcoma by targeting CD44 or EGFR

doi: 10.1016/j.tranon.2024.101966

Figure Lengend Snippet: Concentration and exposure-dependence and target-specific cell death of CD44-IR700 and EGFR-IR700. CD44-IR700-mediated phototoxicity was dependent on the concentration of CD44-IR700 (a) or exposure dose (b). (c) CD44-specific cell death only occurred when VA-ES-BJ cells were exposed to CD44-IR700 and light irradiation. EGFR-IR700-mediated phototoxicity was dependent on the concentration of EGFR-IR700 (d) or exposure dose (e). (f) EGFR-specific cell death only occurred when VA-ES-BJ cells were exposed to both EGFR-IR700 and light irradiation.

Techniques: Concentration Assay, Irradiation

Journal: Translational Oncology

Article Title: Photoimmunotheranostics of epithelioid sarcoma by targeting CD44 or EGFR

doi: 10.1016/j.tranon.2024.101966

Figure Lengend Snippet: Preferential accumulation of CD44-IR700 or EGFR-IR700 in VA-ES-BJ tumors. NIR fluorescence imaging of nude mice bearing bilateral VA-ES-BJ tumors over a 24-h period. 100 μg of CD44-IR700 (a) or EGFR-IR700 (b) or IgG-IR700 (c) antibody conjugate was injected i.v. (d) Plots of tumor to normal (T/N) ratios measured at 0 min, 1 h, 6 h and 24 h p.i. ROIs of tumors and normal tissue were drawn on in vivo NIR fluorescence images. T/N ratios were derived from the MFI of ROIs and represented as Mean ± SD for all three groups ( n = 10 per group). Comparisons of T/N ratios between the groups at 6 h p.i. and 24 h p.i. are shown on the right, ns: no significance, * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. (e) Ex vivo fluorescence images of whole tumors resected at 24 h p.i. (f) The MFI of resected tumors at 24 h p.i. in different groups. Values represent Mean ± SD from six tumors per group ( n = 6). (g) Bio-distribution of antibody conjugates in main organs and tumors resected at 24 h p.i. H, heart; Li, liver; Sp, spleen; Lu, lung; K, kidney; In, intestine; St, stomach; Mu, muscle; Bn, bone; Bl, blood; T, tumor. Values (Mean ± SD) are normalized to % injected dose/g (% ID /g) from three mice per group ( n = 3). Only comparisons with p value less than 0.05 displayed, * P < 0.05, *** P < 0.001, **** P < 0.0001.

Techniques: Fluorescence, Imaging, Injection, In Vivo, Derivative Assay, Ex Vivo

Journal: Translational Oncology

Article Title: Photoimmunotheranostics of epithelioid sarcoma by targeting CD44 or EGFR

doi: 10.1016/j.tranon.2024.101966

Figure Lengend Snippet: In vivo CD44-IR700 or EGFR-IR700-PIT. (a) Growth curve of VA-ES-BJ tumors. 100 μg of CD44-IR700 or EGFR-IR700 or IgG-IR700 was injected i.v. on day 0 and on day 7. All the groups were monitored for three weeks after injection except for two weeks in PBS group. PIT groups received light exposure at 200 J/cm 2 , 24 h after each injection (pointed by blue arrows) and mice in the no PIT groups were shielded from light. Values represent Mean ± SEM from at least four mice per group ( n ≥ 4). (b) The comparisons of tumor volumes between the groups at each monitoring point, ns: no significance, * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001. (c) Photographs of representative VA-ES-BJ tumors harvested at the end of treatment.

Techniques: In Vivo, Injection

Journal: Translational Oncology

Article Title: Photoimmunotheranostics of epithelioid sarcoma by targeting CD44 or EGFR

doi: 10.1016/j.tranon.2024.101966

Figure Lengend Snippet: Short-term effects of PIT on CD44 and EGFR. (a) Immunoblots probing CD44 and EGFR protein in VA-ES-BJ tumors. (b) Representative flow cytometry profiles of CD44 and EGFR expression in tumors after receiving PITs. 100 μg of CD44-IR700 or EGFR-IR700 or 100 μl of PBS was injected i.v. PIT groups received light exposure at 200 J/cm 2 24 h after injection and mice in the PBS group were shielded from light. VA-ES-BJ tumors were harvested three days after single PIT. Tumors were dissociated into a single cell suspension and only live cells were included in the flow cytometry analysis.

Techniques: Western Blot, Flow Cytometry, Expressing, Injection, Suspension

Journal: Translational Oncology

Article Title: Photoimmunotheranostics of epithelioid sarcoma by targeting CD44 or EGFR

doi: 10.1016/j.tranon.2024.101966

Article Snippet: Anti-CD44 monoclonal rat IgG2b antibody (clone IM7), rat IgG2b isotype control (clone LTF-2), anti-MUC1monoclonal mouse IgG3 antibody (clone C595), and anti-EGFR monoclonal mouse IgG1 antibody (clone 225) were purchased from BioXcell (Lebanon, New Hampshire, USA).

Techniques: Over Expression, Expressing, Flow Cytometry, Staining, Immunostaining

Journal: Translational Oncology

Article Title: Photoimmunotheranostics of epithelioid sarcoma by targeting CD44 or EGFR

doi: 10.1016/j.tranon.2024.101966

Techniques: Concentration Assay, Irradiation

Journal: Translational Oncology

Article Title: Photoimmunotheranostics of epithelioid sarcoma by targeting CD44 or EGFR

doi: 10.1016/j.tranon.2024.101966

Techniques: Fluorescence, Imaging, Injection, In Vivo, Derivative Assay, Ex Vivo

Journal: Translational Oncology

Article Title: Photoimmunotheranostics of epithelioid sarcoma by targeting CD44 or EGFR

doi: 10.1016/j.tranon.2024.101966

Techniques: In Vivo, Injection

Journal: Translational Oncology

Article Title: Photoimmunotheranostics of epithelioid sarcoma by targeting CD44 or EGFR

doi: 10.1016/j.tranon.2024.101966

Techniques: Western Blot, Flow Cytometry, Expressing, Injection, Suspension

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