Journal: International Journal of Molecular Sciences

Article Title: HSPA4 Is a Biomarker of Placenta Accreta and Enhances the Angiogenesis Ability of Vessel Endothelial Cells

doi: 10.3390/ijms23105682

Figure Lengend Snippet: Details of the differentially abundant proteins identified between five NP and five PAS placenta tissues. We used 2D-DIGE and MALDI TOF/TOF to quantify and identify proteins, respectively. The theoretical pI/Mr, sequence coverage and score were determined by the MALDI TOP/TOF assay. The fold change and p -value were calculate by the 2D-DIGE assay. Positive fold change denoted a higher abundance in PAS samples. p -value was calculated based on a t -test.

Article Snippet: Finally, tissue sections were applied with 100 μL diluted primary HSPA4 antibody (1:200, Cat. No. HPA010023, MERK) and incubated at 4 °C, overnight, followed by a PBS wash. Next, the tissue sections were in order subjected to incubation with 100 μL appropriately ImmPRESSTM HRP REAGENT KIT (Cat. No. MP-7500, VECTOR) at room temperature for 30 min and with ImmPACTTMDAB (Cat. No. SK-4105, VECTOR) for 3 min. Tissue sections were further counterstained by immersing sides in Hematoxylin for 1 min, followed by running tap water rinse for 10 min.

Techniques: Sequencing

Journal: International Journal of Molecular Sciences

Article Title: HSPA4 Is a Biomarker of Placenta Accreta and Enhances the Angiogenesis Ability of Vessel Endothelial Cells

doi: 10.3390/ijms23105682

Figure Lengend Snippet: The results of protein quantification with 2D-DIGE. After labeling, the images of gels were scanned in a Typhoon 9400 scanner and further analyzed with Decyder software to quantify protein abundance. HSPA4 and PRKAR2 had higher abundances in PAS tissues, whereas CSH was more abundant in NP tissues.

Article Snippet: Finally, tissue sections were applied with 100 μL diluted primary HSPA4 antibody (1:200, Cat. No. HPA010023, MERK) and incubated at 4 °C, overnight, followed by a PBS wash. Next, the tissue sections were in order subjected to incubation with 100 μL appropriately ImmPRESSTM HRP REAGENT KIT (Cat. No. MP-7500, VECTOR) at room temperature for 30 min and with ImmPACTTMDAB (Cat. No. SK-4105, VECTOR) for 3 min. Tissue sections were further counterstained by immersing sides in Hematoxylin for 1 min, followed by running tap water rinse for 10 min.

Techniques: Labeling, Software, Quantitative Proteomics

Journal: International Journal of Molecular Sciences

Article Title: HSPA4 Is a Biomarker of Placenta Accreta and Enhances the Angiogenesis Ability of Vessel Endothelial Cells

doi: 10.3390/ijms23105682

Figure Lengend Snippet: The results of western blot assays. We used western blot assays to validate the variations in protein abundance determined with 2D-DIGE. ( a ) The western blot results. ( b ) With GAPDH as an internal control, HSPA4 and CSH reached statistical significance ( n = 5). * denoted p -value < 0.05 ( t -test). Data was presented as the mean ± S.D.

Article Snippet: Finally, tissue sections were applied with 100 μL diluted primary HSPA4 antibody (1:200, Cat. No. HPA010023, MERK) and incubated at 4 °C, overnight, followed by a PBS wash. Next, the tissue sections were in order subjected to incubation with 100 μL appropriately ImmPRESSTM HRP REAGENT KIT (Cat. No. MP-7500, VECTOR) at room temperature for 30 min and with ImmPACTTMDAB (Cat. No. SK-4105, VECTOR) for 3 min. Tissue sections were further counterstained by immersing sides in Hematoxylin for 1 min, followed by running tap water rinse for 10 min.

Techniques: Western Blot, Quantitative Proteomics, Control

Journal: International Journal of Molecular Sciences

Article Title: HSPA4 Is a Biomarker of Placenta Accreta and Enhances the Angiogenesis Ability of Vessel Endothelial Cells

doi: 10.3390/ijms23105682

Figure Lengend Snippet: The results of IHC assays. We used IHC assay to examine the abundance of HSPA4 protein among the FFPE trophoblast villous. The scanned images were analyzed with GraphPad Prism 5. ( a ) IHC result on one of the trophoblast villous from one PAS and one normal pregnant subject. ( b ) Quantification result of the placenta samples from five PAS and five normal pregnant subjects. * denoted p -value < 0.05.

Article Snippet: Finally, tissue sections were applied with 100 μL diluted primary HSPA4 antibody (1:200, Cat. No. HPA010023, MERK) and incubated at 4 °C, overnight, followed by a PBS wash. Next, the tissue sections were in order subjected to incubation with 100 μL appropriately ImmPRESSTM HRP REAGENT KIT (Cat. No. MP-7500, VECTOR) at room temperature for 30 min and with ImmPACTTMDAB (Cat. No. SK-4105, VECTOR) for 3 min. Tissue sections were further counterstained by immersing sides in Hematoxylin for 1 min, followed by running tap water rinse for 10 min.

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: HSPA4 Is a Biomarker of Placenta Accreta and Enhances the Angiogenesis Ability of Vessel Endothelial Cells

doi: 10.3390/ijms23105682

Figure Lengend Snippet: The results of the tube formation assay. We applied a tube formation assay to investigate whether HSPA4 promoted the angiogenesis ability of HUVECs. The cell growth morphologies of HUVECs transfected with empty pcDNA3.1 construct ( a , control set) or with HSPA4 expression construct ( b , HSPA4 set). ( c , d ) AngioTool analyzed the cell growth morphology and marked the vessels (the thick red lines), highlighted the junctions (the light blue points), and depicted the outlines of vessels (the thin orange lines). ( e ) By analyzing 12 pictures from four independent assays (four independent assays/wells * three pictures), we evaluated the angiogenesis ability by comparing these values. For simplicity, the values in the control set were normalized as one. Data was presented as the mean ± S.D. *** and **** denoted p -value < 0.001 and p -value < 0.0001, respectively.

Article Snippet: Finally, tissue sections were applied with 100 μL diluted primary HSPA4 antibody (1:200, Cat. No. HPA010023, MERK) and incubated at 4 °C, overnight, followed by a PBS wash. Next, the tissue sections were in order subjected to incubation with 100 μL appropriately ImmPRESSTM HRP REAGENT KIT (Cat. No. MP-7500, VECTOR) at room temperature for 30 min and with ImmPACTTMDAB (Cat. No. SK-4105, VECTOR) for 3 min. Tissue sections were further counterstained by immersing sides in Hematoxylin for 1 min, followed by running tap water rinse for 10 min.

Techniques: Tube Formation Assay, Transfection, Construct, Control, Expressing

Journal: Cancers

Article Title: Reticulocalbin 3 Is a Novel Mediator of Glioblastoma Progression.

doi: 10.3390/cancers15072008

Figure Lengend Snippet: Figure 5. RCN3 knockdown enhances the survival of tumor-bearing mice. (A) GSC 040815 cells that stably express control (n = 8) or RCN3 (n = 7) shRNA were implanted intracranially into NOD-SCID mice, and survival was plotted using Kaplan–Meier curves. **** p < 0.0001 by log-rank test. (B) Tumor tissues collected from control and RCN3 knockdown groups were subjected to IHC staining for Ki67 (nuclear), RCN3 (cytoplasmic), and CD44 (membrane), and staining intensity was quantitated. *** p < 0.001, **** p < 0.0001 by student’s t-test.

Article Snippet: Briefly, tumor sections were incubated with RCN3 antibody (Abcam Cat# ab204178—1:500 dilution), Ki67 antibody (Abcam Cat# ab16667—1:100 dilution), or CD44 antibody (Cell Signaling Cat# 37259S—1:200 dilution) overnight, followed by secondary antibody incubation for 30 min (Vector Labs Cat# MP-7500).

Techniques: Knockdown, Stable Transfection, Control, shRNA, Immunohistochemistry, Membrane, Staining