Journal: Scientific Reports

Article Title: Cancer derived peptide of vacuolar ATPase ‘a2’ isoform promotes neutrophil migration by autocrine secretion of IL-8

doi: 10.1038/srep36865

Figure Lengend Snippet: Immunofluorescent staining of ( a ), phosphorylated FAK (pY397) or ( b ), phosphorylated Src (pY416) was performed in neutrophils using confocal microscopy. Neutrophils treated with either PBS (vehicle control) or a2NTD were plated on poly Lysine coated 8 well chamber slides for thirty minutes at 37 °C CO 2 incubator. Cells were fixed, permeabilized, and stained with anti-pFAK (pY397) or anti-pSrc (pY416) primary antibody, followed by secondary antibody AF594 (red), Dapi (blue). Representative images were presented from 3 different experiments. Original maginification 600x; scale bars; 10 μm. Quantification of the intensity of the immunofluorescent staining of ( c ), pFAK (pY397) or ( d ), pSrc (pY416) in neutrophils was performed using FV10i Fluoview Ver.3.0 software. Data are shown as relative average fluorescence intensity to control ± SEM. ***P < 0.001, as compared with control neutrophils.

Article Snippet: After thirty minutes incubation at 37 °C, CO 2 incubator, cells were fixed with 4% paraformaldehyde, permeabilized by alcohol/acetone, blocked and stained with anti-human FAK (BD Biosciences), anti-human FAK (PY397) (BD Biosciences) or anti-human Src (PY416) (Cell Signaling) for one hour at room temperature.

Techniques: Staining, Confocal Microscopy, Control, Software, Fluorescence

Journal: Scientific Reports

Article Title: Bioactive glasses promote rapid pre-osteoblastic cell migration in contrast to hydroxyapatite, while carbonated apatite shows migration inhibiting properties

doi: 10.1038/s41598-023-47883-2

Figure Lengend Snippet: Migration of MC3T3-E1 pre-osteoblastic cells in response to different biomaterials and the expression levels of migration related proteins. ( a ) Number of migrated cells in the Boyden chamber assay at different time points. Results are shown as normalized to control within each time point. ( b – d ) Representative cropped images of the protein expression levels of FAK and SRC ( b ), pY576-FAK and pY529-SRC ( c ), and pY416-SRC ( d ) as detected by Western blot. Each figure ( b – d ) is from a separate membrane. Full membranes are available in Supplementary figure S4. ( e – i ) Quantitative analysis of protein bands is displayed for SRC ( e ), pY529-SRC ( f ), pY416-SRC ( g ), FAK ( h ), and pY576-FAK ( i ), where the control group was assigned as value of 1 within each blot. All data are presented as the mean ± standard deviation of three independent experiments. Parametric data was analyzed using unpaired t-test with Welch’s correction for unequal variances where applicable, and non-parametric data using Mann–Whitney test. Statistical significance is referred to as # p ≤ 0.05 between control and experimental group, and as *p ≤ 0.05, **p ≤ 0.01, and ***p ≤ 0.001 between indicated groups. HAP hydroxyapatite, CAP carbonated apatite, FAK focal adhesion kinase, BMP2 bone morphogenetic protein 2.

Article Snippet: Total protein (15 µg) was run on pre-cast gels (Bio-Rad, 4569033) and electro-transferred to polyvinylidene difluoride (PVDF) membranes, which were blocked with BSA in TBST (5% w/v), followed by incubation with primary antibodies for SRC (Invitrogen, PA5-17717, 1:1000), pY416-SRC (Invitrogen, PA5-97366, 1:1000, pY529-SRC (Invitrogen, 44-662G, 1:1000), FAK (Invitrogen, AHO0502, 1:100) and pY576-FAK (Invitrogen, PA5-104964, 1:750) diluted in TBST.

Techniques: Migration, Expressing, Boyden Chamber Assay, Western Blot, Membrane, Standard Deviation, MANN-WHITNEY