Anti-Lactoferrin/LTF Antibody Picoband®

Western blot analysis of LTF using anti-LTF antibody (A00633-1).
Electrophoresis was performed on a 10% SDS-PAGE gel at 80V (Stacking gel) / 120V (Resolving gel) for 2 hours. The sample well of each lane was loaded with 30 ug of sample under reducing conditions.
Lane 1: human hepatocellular carcinoma paracancerous tissue (HCCP) lysates.
After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% non-fat milk/TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-LTF antigen affinity purified polyclonal antibody (A00633-1) at 0.5 μg/mL overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody (Catalog # BA1054) at a dilution of 1:5000 for 1.5 hour at RT. The signal is developed using an ECL Plus Western Blotting Substrate (Catalog # AR1196-200) with Tanon 5200 system. A specific band was detected for LTF at approximately 85 kDa. The expected band size for LTF is at 78 kDa.

IHC analysis of LTF using anti-LTF antibody (A00633-1).
LTF was detected in a paraffin-embedded section of human prostatic cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2 μg/ml rabbit anti-LTF Antibody (A00633-1) overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Rabbit IgG Super Vision Assay Kit (Catalog # SV0002) with DAB as the chromogen.

LTF, a ferroptosis-related gene, is identified in aging cochleae. (A) The volcano diagram of shows 43385 DEGs between the old group ( ~864316) and young group ( ~864308). (B) The heatmap shows 28 statistically significant DEGs. (C) Functional analysis of the statistically significant DEGs. (D) LTF, the hub gene, is obtained from the intersection of Up-Gens, Down-Gens (based on the statistically significant DEGs), and FRGs. (E) The PPI network shows that 12 proteins are interacting with LTF.
Index in PubMed under a CC BY license. PMID: 38282692

TF-miRNA-mRNA network in cochlear ferroptosis. (A) The volcano diagram of shows 7512 DEMs between the old group ( ~1095954) and young group ( ~1095948). (B) The heatmap shows 12 statistically significant DEMs. (C–F) TFs predicted by mmu-mir-130b (C) , mmu-mir-205 (D) , hsa-mir-130b (E) , and hsa-mir-205 (F) . (G) CEBPA and CEBPB were predicted by mir-130b. (H) STAT3 was predicted by mir-205. (I,J) The top 20 TFs predicted by LTF in mice (I) and humans (J) , CEBPA (Red box) was identified through the intersection of (I,J) , and TFs predicted by miRNAs in (G,H) . (K) The regulatory network was constructed.
Index in PubMed under a CC BY license. PMID: 38282692

Ferroptosis in aging HEI-OC1 cells. (A) SA-β-gal staining in HEI-OC1 cells treated with different concentrations of D-gal for 48 h. (B) Quantification of SA-β-positive cells in (A) . Compared with the control group, the percentage of positive cells is observed to have a statistical difference at 20 mg/mL D-gal ( p < 0.001; N = 3). (C–E) Compared with the control group, the expression of Fe 2+ (C) and MDA (D) is increased in aging HEI-OC1 cells with 20 mg/mL D-gal ( p = 0.006 and p < 0.001; N = 3). Oppositely, the expression of LTF (E) is decreased ( p < 0.001; N = 3). More importantly, Liproxstatin-1 reversed the above phenomenon of ferroptosis in aging HEI-OC1 cells with 20 mg/mL D-gal ( p = 0.038, p = 0.012, and p = 0.046; N = 3). (F) Immunofluorescence staining also shows decreased expression of LTF in aging HEI-OC1 cells with 20 mg/mL D-gal (LTF-green, DAPI-blue). Ctrl, control; Lip, Liproxstatin-1.
Index in PubMed under a CC BY license. PMID: 38282692

Ferroptosis in aging cochlear explants. (A) SA-β-gal staining in the basement membrane treated with different concentrations of D-gal for 48 h. (B) Quantification of SA-β-positive cells in (A) . Compared with the control group, the percentage of positive cells is observed to have a statistical difference at 30 mg/mL D-gal ( p < 0.001; N = 3). (C–E) Compared with the control group, the expression of Fe 2+ (C) and MDA (D) is increased in aging cochlear explants with 30 mg/mL D-gal ( p < 0.001; N = 5). Oppositely, the expression of LTF (E) is decreased in aging cochlear explants with 30 mg/mL D-gal ( p < 0.001; N = 5). OHC, outer hair cell; IHC, inner hair cell; Ctrl, control.
Index in PubMed under a CC BY license. PMID: 38282692

Ferroptosis in aging cochleae. (A) Compared with young mice (2mo), the distinctly increased ABR thresholds are shown in old mice (10mo) at all frequencies ( p < 0.001; N = 6 mice). (B–D) Compared with young mice, the expression of Fe 2+ (B) and MDA (C) is increased in aging cochleae ( p < 0.001; N = 5). Oppositely, the expression of LTF (D) is decreased ( p < 0.005; N = 3).
Index in PubMed under a CC BY license. PMID: 38282692

IF analysis of LTF using anti-LTF antibody (A00633-1).
LTF was detected in a paraffin-embedded section of human prostatic cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 5 μg/mL rabbit anti-LTF Antibody (A00633-1) overnight at 4°C. DyLight®488 Conjugated Goat Anti-Rabbit IgG (BA1127) was used as secondary antibody at 1:500 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Western blot analysis of Lactoferrin/LTF using anti-Lactoferrin/LTF Antibody (A00633-1).
Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30 ug of sample under reducing conditions.
Lane 1: Total LTF from mouse neutrophil cell lysates.
Lane 2: degranulated LTF. Ratio 1:20.
After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% BSA for 1.5 hour at RT. The membrane was incubated with rabbit anti-Lactoferrin/LTF antigen affinity purified polyclonal antibody (A00633-1) at 1:2000 1h overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:3000 dilution at RT for 1 hour. The signal is developed using HRP. A specific band was detected for Lactoferrin/LTF at approximately 77 kDa. The expected band size for Lactoferrin/LTF is at 77 kDa.