Characterization of ζ-associated protein, 70 kd (ZAP70)–deficient human lymphocytes Chaim M. Roifman, MD, Harjit Dadi, PhD, Raz Somech, MD, Amit Nahum,

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Characterization of ζ-associated protein, 70 kd (ZAP70)–deficient human lymphocytes Chaim M. Roifman, MD, Harjit Dadi, PhD, Raz Somech, MD, Amit Nahum, MD, PhD, Nigel Sharfe, PhD Journal of Allergy and Clinical Immunology Volume 126, Issue 6, Pages 1226-1233.e1 (December 2010) DOI: 10.1016/j.jaci.2010.07.029 Copyright © 2010 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 1 TCR repertoire. TCR Vβ families were assessed by means of flow cytometry. Double staining with each Vβ family antibody and either anti-CD4 or anti-CD8 was performed in patient and control samples. Journal of Allergy and Clinical Immunology 2010 126, 1226-1233.e1DOI: (10.1016/j.jaci.2010.07.029) Copyright © 2010 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 2 Quantitative analysis of inflammatory related gene products. Expression of mRNA for the cytokines IL-4, IL-10, and TGF-β; the cell-surface receptors CD8, CTLA-4, and CD5; and the transcription factor GATA-3 were analyzed and compared with that seen in the control sample. Data are expressed as patient fold change over control fold change. Journal of Allergy and Clinical Immunology 2010 126, 1226-1233.e1DOI: (10.1016/j.jaci.2010.07.029) Copyright © 2010 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 3 Analysis of apoptosis in Zap-70−/− peripheral blood and thymus. A, Using microarray analysis of mRNA levels, BCL2 levels are increased, whereas Fas mRNA is reduced relative to that seen in control samples. B, Patient and control cells were incubated with 30 mg/mL anti-CD95 or cisplatin, followed by staining with Annexin V–FITC and analyzed by means of flow cytometry. C, Immunohistochemical terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining of patient and control thymus sections. Apoptotic cells stain dark brown. The number of dark brown–stained apoptotic cells in control samples is more than 4-fold higher than in the patient section. Journal of Allergy and Clinical Immunology 2010 126, 1226-1233.e1DOI: (10.1016/j.jaci.2010.07.029) Copyright © 2010 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Checkpoints of T-cell differentiation Checkpoints of T-cell differentiation. As indicated, the figure represents the developmental stage of thymocytes, the unique cell-surface markers for each stage, and the corresponding TCR rearrangement events during thymocyte maturation. Red dashed lines highlight the putative stage of TCR rearrangement affected by mutations in recombinase activating gene 1, ADA, or ZAP70. Journal of Allergy and Clinical Immunology 2010 126, 1226-1233.e1DOI: (10.1016/j.jaci.2010.07.029) Copyright © 2010 American Academy of Allergy, Asthma & Immunology Terms and Conditions