Journal: Frontiers in Immunology

Article Title: Next-Generation Manufacturing Protocols Enriching T SCM CAR T Cells Can Overcome Disease-Specific T Cell Defects in Cancer Patients

doi: 10.3389/fimmu.2020.01217

Figure Lengend Snippet: Optimized CAR T cell manufacturing is able to rescue the phenotype of T cells derived from cancer patients in a disease-specific manner. (A–E) Phenotypical characterization of healthy donor- (HD), B-cell acute lymphoblastic leukemia- (B-ALL) and pancreatic ductal adenocarcinoma (PDAC)-derived T cells in PB prior activation with the nanomatrix protocol, illustrating (A) CD4/CD8 ratio ( n = 16 for HD; n = 12 for B-ALL; n = 7 for PDAC), (B) T cell memory phenotype ( n = 7 for HD; n = 13 for B-ALL; n = 7 for PDAC), T N : CD45RA + CD62L + CD95 − ; T SCM : CD45RA + CD62L + CD95 + ; T CM : CD45RA − CD62L + ; T EM : CD45RA − CD62L − ; T EMRA : CD45RA + CD62L − , (C) CD127 expression ( n = 16 for HD; n = 13 for B-ALL; n = 7 for PDAC) and (D) PD-1 expression ( n = 16 for HD; n = 13 for B-ALL; n = 7 for PDAC). (E) Analysis of PB T cell exhaustion by means of SPICE algorithm calculated on both CD4 and CD8 compartments, depicting concomitant expression of inhibitory and senescence markers (PD-1, TIM-3 and CD57, n = 7 for HD; n = 11 for B-ALL; n = 7 for PDAC). (F) Fold expansion in culture of HDs and patient-derived T cells after activation with the nanomatrix protocol and transduction with a CD19.BBz CAR ( n = 10 for HD; n = 7 for B-ALL; n = 7 for PDAC). (G–I) Phenotypical characterization of HDs and patient-derived CD19.BBz CAR T cells after activation with the nanomatrix protocol, illustrating (G) the CD4/CD8 ratio, (H) T cell memory phenotype and (I) PD-1 expression ( n = 12 for HD; n = 7 for B-ALL; n = 7 for PDAC). Data are reported as the result of mean ±SEM and paired t -test and two-way ANOVA statistical analyses are reported when statistically significant (* p < 0.05, ** p < 0.01, *** p < 0.001,**** p < 0.0001).

Article Snippet: CAR T cells and mouse samples were stained with one or more of the following conjugated monoclonal antibodies: anti-human CD3 PB (Biolegend, clone HIT3a), CD45 BV510 (Biolegend, clone HI30), CD271 PE-Cy7 (Biolegend, clone CD40-1457), CD271 PE (BD, clone C40-1457), CD4 FITC (Biolegend, clone SK3), CD14 APC (Biolegend, clone M5E2), CD19 APC/Cy7 (Biolegend, clone HIB19), HLA-DR APC/Cy7 (Biolegend, clone L243), CD45RA FITC (Biolegend, clone HI100), CD62L APC (Biolegend, clone DREG-56), CD8 PerCP (BD, clone SK1), CD57 APC/Cy7 (Milteny, clone TB03), CD127 PE (Biolegend, clone A019D5), EGFR PE (Biolegend, clone AY13), CD95 (Fas/APO-1) PE-Cy7 (Biolegend, clone DX2), CD279 (PD-1) PE-Cy7 (Biolegend, clone EH12.1), TIM-3 Alexa Fluor 488 (Biolegend, clone F38-2E2), and anti-mouse CD45 PerCP (Biolegend, clone 30-F11).

Techniques: Derivative Assay, Activation Assay, Expressing, Transduction

Journal: Frontiers in Immunology

Article Title: Next-Generation Manufacturing Protocols Enriching T SCM CAR T Cells Can Overcome Disease-Specific T Cell Defects in Cancer Patients

doi: 10.3389/fimmu.2020.01217

Figure Lengend Snippet: Retrospective correlation of CAR T cell features with therapeutic outcomes in vivo . (A) Bioluminescence detection of Lucia + /LNGFR + NALM-6 systemic growth in NSG mice after treatment with 3 × 10 6 CD19.BBz CAR T cells of HD- and patient-derived origin, retrospectively clustered as short- and long-term responders ( n = 18 for long-term responders, red lines; n = 29 for short-term responders, black lines. More in detail: n = 26 mice for HD, 8 donors, continuous lines; n = 9 mice for B-ALL, 3 patients: #5, #8, #9 in , dashed lines; n = 12 mice for PDAC, 3 patients: #1, #2, #3 in , dotted lines; n = 10 mice for Mock control, 3 donors, gray lines). Data are represented as single lines identifying individual mice treated with CD19.BBz CAR T cells of HD- and patient-derived origin and differently clustered on the basis of CAR T cell anti-leukemia activity. (B) Left panel: analysis of CD19.BBz CAR T cell expansion kinetic (cell count/μL) of short- and long-term responders in the peripheral blood of Lucia + /LNGFR + NALM-6 bearing mice; Right panel: frequency of CD8 + CAR T cells during in vivo expansion. Data are reported as the result of mean ± SEM and two-way ANOVA statistical analyses are reported when statistically significant (* p < 0.05; **** p < 0.0001). (C) Relationships between CAR T cell phenotype and in vivo growth after leukemia encounter, correlating the initial proportion of T SCM , T EM , CD8 + and CD57 + CAR T cells with in vivo expansion levels at day 7 ( n = 12 for T SCM , T EM , CD8 and CD57). R and P -values for each correlation are reported.

Article Snippet: CAR T cells and mouse samples were stained with one or more of the following conjugated monoclonal antibodies: anti-human CD3 PB (Biolegend, clone HIT3a), CD45 BV510 (Biolegend, clone HI30), CD271 PE-Cy7 (Biolegend, clone CD40-1457), CD271 PE (BD, clone C40-1457), CD4 FITC (Biolegend, clone SK3), CD14 APC (Biolegend, clone M5E2), CD19 APC/Cy7 (Biolegend, clone HIB19), HLA-DR APC/Cy7 (Biolegend, clone L243), CD45RA FITC (Biolegend, clone HI100), CD62L APC (Biolegend, clone DREG-56), CD8 PerCP (BD, clone SK1), CD57 APC/Cy7 (Milteny, clone TB03), CD127 PE (Biolegend, clone A019D5), EGFR PE (Biolegend, clone AY13), CD95 (Fas/APO-1) PE-Cy7 (Biolegend, clone DX2), CD279 (PD-1) PE-Cy7 (Biolegend, clone EH12.1), TIM-3 Alexa Fluor 488 (Biolegend, clone F38-2E2), and anti-mouse CD45 PerCP (Biolegend, clone 30-F11).

Techniques: In Vivo, Derivative Assay, Control, Activity Assay, Cell Counting

Journal: Cell

Article Title: Spatial proteogenomics reveals distinct and evolutionarily conserved hepatic macrophage niches

doi: 10.1016/j.cell.2021.12.018

Figure Lengend Snippet:

Article Snippet: Anti-Human CD74 (REA1103) PE , Miltenyi Biotec , 130-119-203; RRID: AB_2733836.

Techniques: Purification, Recombinant, Staining, cDNA Synthesis, Gene Expression, Software, Microscopy

Journal: bioRxiv

Article Title: Cryo-EM of a Divergent Herpesvirus Reveals Structural Conservation and Novelty Including a Portal-Vertex Tegument Protein with Multiple Macrodomain-like Folds

doi: 10.64898/2026.02.13.705735

Figure Lengend Snippet: (A) Terminase based phylogeny of the Alloherpesviridae and related viruses. Amino acid sequence data for selected viruses representing the phylogenetic spread of herpesviruses were used along with that of the bacteriophage T4. The names of virus genera and families are in red and blue font, respectively. The nomenclature in black font consists of the underlying GenBank nucleotide accession number followed by the virus name, which is either a systematic one for classified members of the Alloherpesviridae , or a common one for unclassified viruses grouping in the Alloherpesviridae , and viruses in other families. The names of unclassified viruses are marked with asterisks. The scale shows the number of substitutions per position. (B,C) Micrographs of purified IcHV-1 capsids (B) and virions (C), scale bar = 100nm. (D,E) Icosahedral reconstructions of IcHV-1 capsid (D) and virion (E). Radial-depth cued isosurface representations are shown, viewed along a two-fold symmetry axis. The colour key indicates the radial distance in nm from the particle centre. The penton, hexons, and triplexes in the asymmetric unit are colour-coded and indicated in (D). Two copies of the inner-tegument protein pORF65 are indicated by purple/pink arrows in (E). In the virion reconstruction, density was found in the distal hexon channels, occluding the opening. Cross-sections through the reconstruction showed weak, blurred density radiating from this feature (inset, E).

Article Snippet: IcHV-1 strain Auburn 1 (ATCC VR-665) was propagated in brown bullhead (BB) cells (ATCC CCL-59) in 30 T-175 flasks.

Techniques: Sequencing, Virus, Purification

Journal: bioRxiv

Article Title: Cryo-EM of a Divergent Herpesvirus Reveals Structural Conservation and Novelty Including a Portal-Vertex Tegument Protein with Multiple Macrodomain-like Folds

doi: 10.64898/2026.02.13.705735

Figure Lengend Snippet: Solvent-excluded surface (SES) representations of the asymmetric units of the IcHV-1 capsid (A) and virion (B). The asymmetric units contain 16 copies of the major capsid protein pORF39, (blue): six (P1-P6) make up a complete P-hexon, six (C1-C6) make up a complete C-hexon, and three (E1-E3) contribute to half of an E-hexon. One copy is present and contributed to the penton. The asymmetric unit comprises six triplexes each composed of one copy of pORF53 (purple), and two copies of pORF27 (white). In the virion, two copies of the inner-tegument protein pORF65 are positioned on the Tb and Tc triplexes. Additionally, three copies of pORF42 occlude the distal tips of the hexon-channels. (C) Schematic representation of the virion asymmetric unit coloured as in (A)–(B). (D) A ribbon diagram of the penton pORF39 is shown, color-coded to highlight the domain structure previously defined for orthoherpesvirus MCPs. The IcHV-1 MCP features an extended C-terminal region (red), which folds toward the helix–hairpin domain to form a helical bundle. This region and adopts different conformations in penton versus hexon environments (compare E and F). (E) A ribbon diagram of the penton MCP is shown in rainbow colour-scheme from N-terminus (blue) to C-terminus (red). (F) A ribbon diagram of the hexon MCP is shown in the same rainbow colour-scheme. A second, neighbouring copy is shown in both ribbon and transparent sovent-excluded surface (blue), to highlight the domain insertion of the C-terminal domain in this form of the IcHV-1 capsomere. (G) A ribbon diagram of pORF53 is shown in rainbow colour-scheme, with the associated two copies of pORF27 shown as SES (white). (H) Two copies of pORF27 are shown as ribbon diagrams, one in rainbow colour-scheme and one in white. The associated copy of pORF53 is displayed as a purple SES. (I) SES representation of a P-hexon (blue/gold) and adjacent Tc triplex (purple/white). pORF65 is shown as a ribbon diagram in rainbow colour-scheme.

Article Snippet: IcHV-1 strain Auburn 1 (ATCC VR-665) was propagated in brown bullhead (BB) cells (ATCC CCL-59) in 30 T-175 flasks.

Techniques: Solvent

Journal: bioRxiv

Article Title: Cryo-EM of a Divergent Herpesvirus Reveals Structural Conservation and Novelty Including a Portal-Vertex Tegument Protein with Multiple Macrodomain-like Folds

doi: 10.64898/2026.02.13.705735

Figure Lengend Snippet: (A–C) Ribbon diagram representations of the portal complexes of the IcHV-1 capsid (A), virion (B), and that of HSV-1 (C), each shown in rainbow colouring. The IcHV-1 portal consists of 12 copies of pORF37, likewise the HSV-1 portal complex comprises 12 copies of pUL6. All structures exhibit C12 symmetry. (D, E) Corresponding cryo-EM density maps of the IcHV-1 capsid (D) and virion (E), portals, resolved to 4.3 Å and 4.6 Å respectively. In the capsid map, an additional density is observed on the outer face of the wing domain (purple), assigned by ModelAngelo as the N-terminus of pORF37. Due to discontinuity in the density, we were unable to determine whether this short α-helix originates from the same monomer or is contributed by a neighbouring subunit. (F, G) Structural comparison of HSV-1 pUL6 (F) and IcHV-1 Orf37 (G). Each monomer is shown as a ribbon diagram coloured to highlight the domain structure (left) and rainbow representation (right). (H) Two adjacent virion portal pORF37 monomers are shown as ribbon diagrams (yellow and blue) within the reconstructed volume (transparent surface), to highlight two instances of β-augmentation. Yellow and blue boxes indicate unique monomer–monomer interaction regions. (I) Enlarged view of the yellow-boxed region, showing β-strand augmentation in which a three-stranded β-sheet is assembled by two neighbouring protomers to form the clip region. (J, K) View of the top of the wing domain in the capsid (J) and virion (K) portal, highlighting a rearrangement between the two forms whereby the N-terminal β-strand which contributes to a four-stranded β-sheet in one monomer (black arrows) inserts instead into the same β-sheet in the neighbouring protomer, forming an inter-subunit β-sheet extension. This rearrangement likely accounts for the loss of the N-terminal helical density at the base of the wing domain in the virion portal.

Article Snippet: IcHV-1 strain Auburn 1 (ATCC VR-665) was propagated in brown bullhead (BB) cells (ATCC CCL-59) in 30 T-175 flasks.

Techniques: Cryo-EM Sample Prep, Comparison