fluorescein isothiocyanate fitc coupled concanavalin a lectin  (Vector Laboratories)

Journal: Molecular Therapy. Methods & Clinical Development

Article Title: Mannose-coupled AAV2: A second-generation AAV vector for increased retinal gene therapy efficiency

doi: 10.1016/j.omtm.2024.101187

Figure Lengend Snippet: First covalent coupling of 5 onto the capsid of AAV2 via primary amino groups (A) 1.4 E 12 vg of AAV2-GFP vectors were added to a solution of compound 5 (3 E 5 or 3 E 6 eq) in TBS buffer (pH 9.3) and incubated for 4 h at RT. The same experimental procedure was followed with compound 7 (3 E 6 eq) in TBS at pH 9.3 as a control. (B and C). 5 E 8 vg of the samples were analyzed by western blot using a polyclonal antibody against the capsid to detect VP proteins (B) or using an FITC-concanavalin A lectin (C) to detect mannose sugar. Note that compound 7 is a negative control lacking the Ar-NCS reactive function. (D) 1 E 10 vg of each condition were analyzed by silver nitrate staining. VP1, VP2, and VP3 are the three proteins constituting the AAV capsid. Capsid protein molecular weight is indicated at the right of the images according to a protein ladder.

Article Snippet: FITC-concanavalin A lectin was purchased from Vector Laboratories.

Techniques: Incubation, Western Blot, Negative Control, Staining, Molecular Weight

Journal: Science bulletin

Article Title: Ethanolamine as a biomarker and biomarker-based therapy for diabetic retinopathy in glucose-well-controlled diabetic patients.

doi: 10.1016/j.scib.2023.12.053

Figure Lengend Snippet: Fig. 6. Ethanolamine prevents retinal inflammation in diabetic rats with suppressed microglial activation and leukocyte adhesion. (a) Schematic illustration of the experimental design of ethanolamine gavage intervention in STZ-induced diabetic SD rats for 36 d. (b) Metabolism analysis revealed that total ethanolamine was significantly decreased in diabetic mouse retinas 8 weeks after diabetes induction. Data are expressed as the mean ± SD (n = 3 retinas from distinct mice). (c, d) Fasting plasma glucose levels and body weight were measured weekly after STZ or PBS injection in SD rats with or without ethanolamine gavage intervention (100 mg/kg; n = 24). (e) Ethanolamine inhibited IL-1b expression at the mRNA level in diabetic rat retinas 36 d after DR induction, as determined via qRT-PCR analysis. Data are expressed as the mean ± SEM (n = 3– 6 retinas from distinct rats). (f) Ethanolamine prevented leukocyte adhesion in diabetic rat retinas 30 d after DR induction. Representative images of adherent leukocytes within the retinal vasculature in flat-mounted retinas by lectin staining. Adherent leukocytes are indicated by white arrowheads. Scale bar, 50 lm. (g) Quantification of retinal adherent leukocytes. Data are presented as the mean ± SD. (n = 2–4 retinas from distinct rats). (h) Ethanolamine-mediated inhibition of microglia activation in diabetic rat retinas. Representative retinal cross-sections showed the morphological changes in microglial immunolabeled for IBA-1 (green) in retinas from normal rats and diabetic rats 36 d after DR induction with or without ethanolamine treatment. Scale bar, 25 lm. STZ, diabetes group; STZ + ETN, diabetes with ethanolamine treatment group; Control, normal control group. # P < 0.05 vs. the control group. ## P < 0.01 vs. the control group. ###P < 0.001 vs. the control group. * P < 0.05 vs. the STZ group. ** P < 0.01 vs. the STZ group, and ns indicates non-significant vs. the STZ group.

Article Snippet: Subsequently, to label the adherent leukocytes and vascular endothelium, we perfused 20 lg/mL FITC-coupled concanavalin A lectin (ConA) (FL-1001; Vector Laboratories, USA) in PBS at a dose of 5 mg/kg.

Techniques: Activation Assay, Clinical Proteomics, Injection, Expressing, Quantitative RT-PCR, Staining, Inhibition, Immunolabeling, Control

Journal: The FASEB Journal

Article Title: In contrast to Western diet, a plant-based, high-fat, low-sugar diet does not exacerbate retinal endothelial injury in streptozotocin-induced diabetes

doi: 10.1096/fj.201900462R

Figure Lengend Snippet: Retinal leukocyte accumulation and endothelial injury. Normal and STZ-induced diabetic rats fed regular chow or WD for 2 wk were perfused with FITC-conjugated ConA. Retinas were harvested and flat mounted for histologic analysis. To illustrate a larger region of the retinal flat mount at higher resolution, composite micrographs were generated by merging the digital images from adjacent regions of the retina in a mosaic fashion. A) Representative composite micrographs of accumulated leukocytes (arrowheads) in normal and diabetic animals. Scale bar, 200 µm. B) Quantitation of the number of accumulated leukocytes per retina in normal and diabetic animals on standard chow or WD; n = 7–9 in each group. C) To illustrate endothelial injury, in vivo PI staining (red) was performed. Representative micrographs illustrate injured endothelial cells (arrows) in normal and diabetic animals on standard chow or WD 2 wk after diabetes induction. Scale bar, 200 µm. D) Quantitation of the number of PI-stained endothelial cells per entire retina in normal and diabetic animals on standard chow or WD 2 wk after diabetes induction. *P < 0.05, **P < 0.01 (n = 10–16 in each group).

Article Snippet: Leukocyte adhesion in the retinal microvessels Retinal vessels and adhering leukocytes in control and diabetic animals were labeled with FITC-conjugated concanavalin A (ConA) lectin (Vector Laboratories, Burlingame, CA, USA).

Techniques: Generated, Quantitation Assay, In Vivo, Staining

Journal: The FASEB Journal

Article Title: In contrast to Western diet, a plant-based, high-fat, low-sugar diet does not exacerbate retinal endothelial injury in streptozotocin-induced diabetes

doi: 10.1096/fj.201900462R

Figure Lengend Snippet: Retinal leukostasis and endothelial damage in animals on semipurified diets. Normal and STZ-induced diabetic animals were fed the indicated semipurified diets for 2 wk. The diets were designed to be high in fats, mimicking the composition of plant-based and animal-based sources with and without added cholesterol (0.1% of the total diet). At the end of the period, animals were perfused with FITC-conjugated ConA lectin; subsequently, retinal flat mounts were generated for histologic examinations. A) Quantitation of the number of firmly adhering leukocytes. B) Composite micrographs of representative sections of retinas from indicated groups. C) PI-stained injured or dead endothelial cells per retina in STZ-induced diabetic rats and nondiabetic controls fed indicated diets. D) Representative micrographs showing PI-positive cells in retinal flat mounts. Scale bar, 200 µm. *P < 0.05 (n = 7–14).

Article Snippet: Leukocyte adhesion in the retinal microvessels Retinal vessels and adhering leukocytes in control and diabetic animals were labeled with FITC-conjugated concanavalin A (ConA) lectin (Vector Laboratories, Burlingame, CA, USA).

Techniques: Generated, Quantitation Assay, Staining