rea1112  (Miltenyi Biotec)

Journal: Analytical Chemistry

Article Title: Immunomagnetic Sample Preparation Targeting Host- and Viral-Derived Antigens for HIV‑1 Isolation from Limited Plasma Volumes

doi: 10.1021/acs.analchem.5c06176

Figure Lengend Snippet: Immunoconjugate-mediated capture of HIV-1 from clinical specimens. Five specimens with viral loads ranging from 107,024 to 1,393,440 copies/mL were used. n = 2–3 technical replicates/specimen and n = 3–5 clinical specimens/conjugate, except gp120, gp41, CD45, CD86, ICAM-1, and CD11a/CD18 LFA-1 ( n = 1 specimen) were used. A. % Captured HIV-1 by immunoconjugates incubated with HIV-positive plasma. Healthy human plasma acted as a negative control. Beads or isotype controls show nonspecific capture of HIV-1. B. Captured HIV-1 RNA copies by immunoconjugates incubated with 25 μL patient specimens. C. Captured HIV-1 RNA copies by CD46 immunoconjugate incubated with increasing volumes of HIV-positive plasma (ID #1654, VL: 384,000 copies/mL), ranging from 2 to 50 μL ( n = 2–3 technical replicates). D. % Captured HIV-1 by the combination of CD46 and CD44 immunoconjugate incubated with patient plasma (ID #1654, VL: 384,000 copies/mL). CD44 (1.6 mg) and CD46 (1.6 mg) were used alone or in combination (0.8 mg each, total 1.6 mg). The no capture control shows the input copies present in 25 μL of HIV-positive human plasma, measured by RT-qPCR. The isotype control shows nonspecific capture by IgG isotype armed microbeads. No-capture control served as the baseline HIV RNA copies in the different plasma volumes ( n = 1–3 replicates).

Article Snippet: Mouse IgG antibodies: CD8 (UCH-T4), Cat # sc-1181, Santa Cruz Biotechnology, Inc.; CD18 (Integrin beta-2), Clone ID H52-s, DSHB (Developmental Studies Hybridoma Bank); HIV1gp120, Cat # sc-57810, Santa Cruz Biotechnology, Inc.; CD26, Cat # 555435, BD Pharmingen; integrin alpha-4, Clone ID P4G9, DSHB; CD43 (DF-T1), Cat # sc-6256, Santa Cruz Biotechnology, Inc.; CD46 (1E3D1), Cat # MA5-29113, Invitrogen; β 2 -microglobulin (B2M-01), Cat # MA1–19141, Invitrogen; integrin alpha-L (CD11a), Clone ID MHM.24, DSHB; CD86 (B7-2), Clone ID IT2.2, Invitrogen; CD44, Clone ID H4C4, DSHB; ICAM1, Clone ID P2A4, DSHB; CD45, Clone ID H5A5, DSHB; HLA-DR, Clone ID LN3, Cat # 14-9956-82, Thermo Fisher Scientific; Tim-4 (human):Fc (mouse) (rec.), Cat # CHI-HF-211T4-C100, AdipoGen Life Sciences.

Techniques: Incubation, Clinical Proteomics, Negative Control, Control, Quantitative RT-PCR

Journal: MethodsX

Article Title: Feline leukocyte immunophenotyping: an optimised whole-blood flow cytometry protocol

doi: 10.1016/j.mex.2026.103869

Figure Lengend Snippet: Representative leukocyte-gated histograms illustrating antibody titration for feline leukocyte immunophenotyping. All histograms display singlet leukocytes, defined by FSC-A versus SSC-A morphological gating, followed by FSC—H versus FSC-A singlet discrimination. Titration was performed for CD18, CD21, CD45R, CD4, CD5 and CD8 monoclonal antibodies using three antibody volumes: 10 µL (1:10 dilution), 5.0 µL (1:20 dilution), 3.0 µL (1:33 dilution) and 1.5 µL (1:66 dilution). Minimal working volumes were selected based on optimal signal-to-noise ratios.

Article Snippet: Step 2 – Leukocytes extracellular staining Materials • EDTA Whole blood sample ± Transfix® • 200 μl pipettes • 100 μl pipettes • 10 μl pipettes • Flow cytometry tubes • Permanent marker • Cytometer tube rack Reagents • Monoclonal antibodies: ○ CD5 Anti-cat – clone FE1.1B11 (BIO-RAD®) ○ CD4 Anti-cat – clone vpg34 (BIO-RAD®) ○ CD8 Anti-cat alpha/beta purified – clone vpg9 (BIO-RAD®) ○ Rat Anti-Mouse IgG1 – clone X56 (BIO-RAD®) ○ CD18 Mouse Anti-Dog – clone CA1.4E9 (BIO-RAD®) ○ CD21 Mouse Anti-Dog – clone CA2.1D6 (BIO-RAD®) ○ CD45R Rat Anti-Mouse – clone RA3–6B2 (BIO-RAD®) • 10x Red blood cells (RBC) lysis buffer solution (BD FACSTM lysing solution) • PBS 1% solution Equipment • Countess TM 3 (Thermo Fisher Scientific, USA) • Freezer • Dark incubation chamber • Timer • Vortex (MX-S®, China) • Centrifuge (model 5810R, Eppendorf®, Germany) • Flow cytometry analyser BD FACSCanto II (Becton Dickinson (BD), San Jose, USA) Methods 1.

Techniques: Titration, Bioprocessing