PerCP/Cy5.5® Conjugation Kit - Lightning-Link®

• Flow Cyt

PubMed

Flow Cytometry - PerCP/Cy5.5® Conjugation Kit - Lightning-Link® (AB102911)

Flow Cytometry - PerCP/Cy5.5 Conjugation Kit- Lightning-Link.

Robinson, Andrew P., et al used PerCP/Cy5.5^® Conjugation Kit - Lightning-Link^® (ab102911) as part of characterizing oligodendroglial populations. They used the kit to conjugate PerCP/Cy5.5^® to Mouse monoclonal anti-NG2 antibody for use in flow cytometry.
SJL/J mice were immunized with PLP139–151 and scored daily for clinical disease. A cohort of SJL/J mice was sacrificed, and spinal cords were analyzed by flow cytometry (n = 5). (A) Cells were distinguished from debris by forward and side scatter then singlet cells were gated. Live cells were gated by dead cell exclusion, and CNS resident cells were identified as CD45− or CD45low. (B) Oligodendroglial cells were defined by double positive staining : A2B5+PDGFRα+ early OPCs, A2B5+NG2+ intermediate OPCs, NG2+O4+ late OPCs, O4+MOG+ pre-myelinating oligodendrocytes, and GALC+MOG+ mature oligodendrocytes.

Image from Robinson, Andrew P., et al., PloS one, 9(9):e107649. doi: 10.1371/journal.pone.0107649. Reproduced under the Creative Commons license https://creativecommons.org/licenses/by/4.0/

• Schematic Diagram

Supplier Data

Schematic Diagram - PerCP/Cy5.5® Conjugation Kit - Lightning-Link® (AB102911)

This illustration demonstrates a general procedure of how Lightning-Link® labeling technology enables the direct labeling of antibodies or proteins.

Simply pipette your antibody or biomolecule of choice into the vial of a lyophilized mixture containing the label of interest and incubate for around 3 hours. Please see the ab102911 protocol booklet for more details.

Learn more about our Lightning-Link® conjugation kits here

• Flow Cyt

PubMed

Flow Cytometry - PerCP/Cy5.5® Conjugation Kit - Lightning-Link® (AB102911)

Okagawa, Tomohiro, et al used PerCP/Cy5.5^® Conjugation Kit - Lightning-Link^® (ab102911) as part of examining the effect on proliferation of bovine leukemia virus (BLV)-specific T cells of the administration of Boch5D2. They used the kit to conjugate PerCP/Cy5.5^® to anti-CD8 antibody, clone CC63, for use in flow cytometry. T-cell proliferation specific for BLV antigen stimulation. Carboxyfluorescein diacetate succinimidyl ester (CFSE)-labeled peripheral blood mononuclear cells were cultured in triplicate with fetal lamb kidney (FLK)-BLV antigen, control FLK antigen (A), or gp51 peptides (0.1 and 1 μg/ml) (B) for 6 days. The percentage of CFSElow cells in CD4+ and CD8+γδTCR− T cells was measured by flow cytometry. CFSElow cells represent cells proliferated during cultivation. Each dot represents the mean of three independent experiments. Significant differences were determined by Dunnett's multiple-comparison test across the time points. *,#,†P < 0.05 versus 0 dpi in each stimulation.

Image from Okagawa, Tomohiro, et al., Front Immunol., 8:650, doi: 10.3389/fimmu.2017.00650. Reproduced under the Creative Commons license https://creativecommons.org/licenses/by/4.0/

• Flow Cyt

PubMed

Flow Cytometry - PerCP/Cy5.5® Conjugation Kit - Lightning-Link® (AB102911)

Nishimori, Asami et al used PerCP/Cy5.5^® Conjugation Kit - Lightning-Link^® (ab102911) as part of examining bovine leukemia virus infection. They used the kit to conjugate PerCP/Cy5.5^® to anti-bovine CD8 antibody for use in flow cytometry.
A BLV-infected cow (#368, Holstein, female, 538 kg, 31 months old) was inoculated with 530 mg (1 mg/kg) of the purified 4G12 intravenously. (A) The proliferation of CD4+ and CD8+ T cells against BLV antigen. Peripheral blood mononuclear cells (PBMCs) isolated from the cow which was inoculated with 4G12 were labeled with carboxyfluorescein diacetate succinimidyl ester (CFSE) and cultured without stimulation (medium) or with the supernatant of FLK or FLK-BLV cells for 6 days. After the cultivation, the proliferation of T cells was immediately analyzed by flow cytometry. A P-value less than 0.05 was considered statistically significant. #, P <0.05 (FLK-BLV, versus day 0; one-way ANOVA followed by Dunnett's test). (B) Changes in PD-L1 occupancy on circulating IgM+ B cells calculated by the binding of 4G12 to bovine PD-L1. The occupancy was estimated as the percentage of the in vivo PD-L1 binding occurred at the total available binding sites. (C) Changes in BLV provirus loads in the cow inoculated with 4G12; the y-axis shows the number of BLV copies included in 50-ng DNA extracts of PBMCs. Data are means ± SEM of at least three replicate experiments.

Image from Nishimori, Asami et al. PloS one vol. 12,4 e0174916. 26 Apr. 2017, doi:10.1371/journal.pone.0174916. Reproduced under the Creative Commons license https://creativecommons.org/licenses/by/4.0/

• Flow Cyt

PubMed

Flow Cytometry - PerCP/Cy5.5® Conjugation Kit - Lightning-Link® (AB102911)

Flow Cytometry - PerCP/Cy5.5 Conjugation Kit - Lightning-Link.

Okagawa, Tomohiro, et al used PerCP/Cy5.5^® Conjugation Kit - Lightning-Link^® (ab102911) as part of examining PD-1 and LAG-3 expression. They used the kit to conjugate PerCP/Cy5.5^® to monoclonal anti-CD3 antibody, clone MM1A, for use in flow cytometry.
Expression of PD-1 and LAG-3 on CD4+ T cells in BLV-infected cattle. A Gating strategy and representative dot plots for expression analyses of PD-1 and LAG-3 on IgM−CD3+CD4+γδTCR− T cells from peripheral blood of BLV-infected cattle (AL and EBL). Values in the quadrants indicate percentages of cells. Percentages of PD-1+LAG-3+CD4+ T cells (B), PD-1+LAG-3−CD4+ T cells (C), and PD-1−LAG-3+CD4+ T cells (D) in CD3+CD4+ T-cell population in peripheral blood from BLV-uninfected (BLV − ; n = 15), AL (n = 22), PL (n = 11), and EBL cattle (n = 7). Bars indicate group median percentage. Significant differences between each group were determined using a Kruskal–Wallis test, where P < 0.05 and P < 0.001, indicated by asterisks (* and ***, respectively).

Image from Okagawa, Tomohiro et al., Vet Res., 49(1):50. doi: 10.1186/s13567-018-0543-9. Reproduced under the Creative Commons license https://creativecommons.org/licenses/by/4.0/

• Conjugation

PubMed

Conjugation - PerCP/Cy5.5® Conjugation Kit - Lightning-Link® (AB102911)

PerCP/Cy5.5® Conjugation Kit - Lightning-Link® labeling anti-canine CD4 antibody for Flow cytometry.

Maekawa N et al. used ab102911 as part of examining a canine chimeric monoclonal antibody targeting PD-L1.

They used the kit to conjugate PerCP/Cy5.5^® to anti-canine CD4 antibody for use in flow cytometry.

To evaluate cell proliferation, nucleotide analogue 5-ethynyl-2′-deoxyuridine (EdU) was added to the medium on day 2, and cells were harvested after incubation for another 2 h. The lymphocyte population was gated by forward scatter and side scatter, and the incorporation of EdU in (c) CD4+ cells was measured by a flow cytometer. Statistical analysis was performed with a Wilcoxon signed rank-sum test.

Image from Maekawa N et al., Sci rep., 7(1):8951. Fig 2.; doi: 10.1038/s41598-017-09444-2. Reproduced under the Creative Commons license https://creativecommons.org/licenses/by/4.0/