Journal: Proteomics

Article Title: Identification, prioritization and evaluation of glycoproteins for aggressive prostate cancer using quantitative glycoproteomics and antibody-based assays on tissue specimens

doi: 10.1002/pmic.201200541

Figure Lengend Snippet: Standard Curve of ELISA assays for protein quantification of cartilage oligomeric matrix protein (COMP), periostin, membrane primary amine oxidase (VAP-1) and cathepsin L. A), dose-response curves for COMP. B), dose-response curve for periostin. C), dose-response curve for VAP-1. D), dose-response curve for cathepsin L.

Article Snippet: Materials Hydrazide resin and sodium periodate were from Bio-Rad (Hercules, CA); sequencing-grade trypsin and TMB reagent were from Promega (Madison, WI); PNGase F was from New England Biolabs (Ipswich, MA); C18 columns were from Waters (Milford, MA); Recombinant protein, capture and detection antibody of human cartilage oligomeric matrix protein (COMP), periostin, cathepsin L, clusterin and galectin-3 binding protein, streptavidin-HRP conjugates, ELISA plates and human VAP-1 Quantikine ELISA Kit were from R&D systems (Minneapolis, MN); All other chemicals were from Sigma-Aldrich (St. Louis, MO).

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Proteome Science

Article Title: Development of reverse phase protein microarrays for the validation of clusterin, a mid-abundant blood biomarker

doi: 10.1186/1477-5956-7-15

Figure Lengend Snippet: Scanned image of fragments of RPPMs showing spotted triplicates of plasma and mouse IgGs probed with (+) or without (-) primary anti-clusterin antibody, and with secondary fluorescently labeled anti-IgG antibody . Pseudo-color scale, dark blue to white corresponds to increasing fluorescence.

Article Snippet: For the measurement of clusterin we used a Human Clusterin ELISA kit (BioVendor, Czech Republic).

Techniques: Labeling, Fluorescence

Journal: Proteome Science

Article Title: Development of reverse phase protein microarrays for the validation of clusterin, a mid-abundant blood biomarker

doi: 10.1186/1477-5956-7-15

Figure Lengend Snippet: Evaluation parameters of RPPMs . (a) Detection of clusterin in plasma samples using monoclonal and (b) polyclonal primary anti-clusterin antibodies. The log 10 average signal intensity for clusterin was plotted against plasma dilution 1/4 to 1/512. (c) Minimum difference in clusterin concentration detected on RPPMs. Twenty different concentrations of recombinant clusterin were spiked in plasma and the average fluorescence intensity of three slides median normalized was plotted against the ten highest clusterin concentrations. The average fluorescence level (▲) with the value of 2 standard deviations (---) for the ten lowest endogenous clusterin concentrations are used as background clusterin level in this experiment. Arrow points to the concentration of spiked clusterin that yielded a signal at least 2SD above background plasma clusterin. (d) Correlation of clusterin levels measured with ELISA and RPPM (r = 0.989). Clusterin was measured in plasma samples spiked with increasing concentrations (0.9–500 μg/ml) of recombinant clusterin.

Article Snippet: For the measurement of clusterin we used a Human Clusterin ELISA kit (BioVendor, Czech Republic).

Techniques: Concentration Assay, Recombinant, Fluorescence, Enzyme-linked Immunosorbent Assay

Journal: Proteome Science

Article Title: Development of reverse phase protein microarrays for the validation of clusterin, a mid-abundant blood biomarker

doi: 10.1186/1477-5956-7-15

Figure Lengend Snippet: Scanned image of slide containing 149 clinical samples spotted in quadruplicate and probed for clusterin .

Article Snippet: For the measurement of clusterin we used a Human Clusterin ELISA kit (BioVendor, Czech Republic).

Techniques:

Journal: Proteome Science

Article Title: Development of reverse phase protein microarrays for the validation of clusterin, a mid-abundant blood biomarker

doi: 10.1186/1477-5956-7-15

Figure Lengend Snippet: Measurement of clinical samples using RPPMs andELISA . (a) The log 10 median clusterin intensity of all clinical samples analysed was plotted. (b) Clusterin levels in serum and CTAD plasma samples taken from the same individual and processed at different time points (0 min to 24 hrs) were measured with ELISA and (c) RPPM. (d) Correlation of RPPM data and ELISA values obtained for all clinical samples. The scaled data of all 16 data points per sample was averaged and then divided by its respective clusterin concentration measured by ELISA to obtain a ratio. For each sample, the log 2 ratio was plotted against its respective clusterin concentration measured by ELISA.

Article Snippet: For the measurement of clusterin we used a Human Clusterin ELISA kit (BioVendor, Czech Republic).

Techniques: Enzyme-linked Immunosorbent Assay, Concentration Assay

Journal: Human reproduction (Oxford, England)

Article Title: Higher clusterin immunolabeling and sperm DNA damage levels in hypertensive men compared with controls.

doi: 10.1093/humrep/des173

Figure Lengend Snippet: Figure 1 Clusterin immunodetection and TUNEL assay before (total) and after (selected) motile spermatozoa swim-up selection, from the same healthy normotensive subject. Swim-up selection drastically reduces the amount of high-clusterin-positive cells as indicated by the very low number of events revealed in the M3 region by flow cytometry (A and B). By immunocytochemistry, normal spermatozoa labeled in the acrosomal region only, possibly corresponding to the flow cytometry M2 region (large dotted square), were observed in the total sperm population, together with morpho- logical abnormal spermatozoa with highly clusterin labeled head and flagellum, possibly corresponding to the M3 region of flow cytometry (small dotted square) (C). Highly labeled cells were not present after swim-up selection of motile spermatozoa (D). Swim-up selection drastically reduces also the amount of TUNEL-positive spermatozoa (E and F) as indicated by the low number of events revealed in the M4 region by flow cytometry. In the upper central insert, a western blot confirming the molecular weight of sperm clusterin bands specifically recognized by the primary antibody used, is visible. Bands at 80 kDa represent the heterodimeric form of the protein, while the single chains migrate at 40 kDa.

Article Snippet: Rabbit polyclonal anti-human clusterin antibody (H-330, #sc-8354; Santa Cruz Biotechnology, Germany), reacting with clusterin a and b chains, was used.

Techniques: Immunodetection, TUNEL Assay, Selection, Cytometry, Immunocytochemistry, Labeling, Western Blot, Molecular Weight

Journal: Human reproduction (Oxford, England)

Article Title: Higher clusterin immunolabeling and sperm DNA damage levels in hypertensive men compared with controls.

doi: 10.1093/humrep/des173

Figure Lengend Snippet: Figure 2 Flow cytometric analysis of sperm clusterin positivity and DNA fragmentation in control and hypertensive subjects (healthy normotensive subjects, n ¼ 25; hypertensive subjects, n ¼ 25). In A and B representative FACS profiles from control and hypertensive subjects are, respectively, shown, while the numbers indicate the mean value of each group studied. (A) Histograms of clusterin expression profile (x-axis: fluorescence intensity; y-axis: number of events). Cells located within the M1 region are clusterin negative. Cells located within the M2 and M3 regions are intermediate (M2) and highly (M3) clusterin positive. (B) Dot plots of spermatozoa double labeled for clusterin and TUNEL assay (x-axis: intensity of TUNEL fluores- cence; y-axis: intensity of clusterin fluorescence). Graphs were divided into four quarters to define the positivity for different staining. LL (low left): spermatozoa with null or medium fluorescence intensity for clusterin and TUNEL negative. UR (upper right): spermatozoa highly positive for both clusterin and TUNEL. UL (upper left): spermatozoa highly positive for clusterin and TUNEL negative; LR (low right): spermatozoa with null or medium fluorescence intensity for clusterin and TUNEL positive. (C) In the histogram, differences for clusterin (M2 and M3), TUNEL and TUNEL/clusterin M3 mean values (% gated) between the control and the hypertensive groups are shown. **P , 0.01.

Article Snippet: Rabbit polyclonal anti-human clusterin antibody (H-330, #sc-8354; Santa Cruz Biotechnology, Germany), reacting with clusterin a and b chains, was used.

Techniques: Control, Expressing, Labeling, TUNEL Assay, Staining

Journal: Human reproduction (Oxford, England)

Article Title: Higher clusterin immunolabeling and sperm DNA damage levels in hypertensive men compared with controls.

doi: 10.1093/humrep/des173

Figure Lengend Snippet: Figure 3 Confocal micrographs of ejaculated human spermatozoa labeled for clusterin (red fluorescence) and for DNA fragmentation (green fluor- escence). Blue fluorescence identifies nuclei (bar ¼ 10 mm). Phase contrast (A) and double immunolabeling for clusterin and TUNEL (B) of the same microscopic field of total sperm population from a hypertensive subject. The arrow indicates two clusterin-positive spermatozoa and the arrowhead points to a clusterin- and TUNEL-negative spermatozoon. Several cells are positive for both clusterin and TUNEL. (C–E): Higher magnification of a total sperm population from a hypertensive subject; C: merged image of the phase contrast with the three fluorescence channels; D: single fluorescent channel for clusterin; E: single fluorescent channel for damaged DNA. The arrows indicate a spermatozoon positive for both clusterin and TUNEL. The arrowheads indicate spermatozoa positive only for clusterin. The frames highlight spermatozoa that are only TUNEL positive. F–H details of a sperm- atozoon showing strong morphological alterations and DNA damage; (F) merged image of the phase contrast microscopy with the three fluorescent channels; (G) single fluorescent channel for clusterin; (H) single fluorescent channel for fragmented DNA.

Article Snippet: Rabbit polyclonal anti-human clusterin antibody (H-330, #sc-8354; Santa Cruz Biotechnology, Germany), reacting with clusterin a and b chains, was used.

Techniques: Labeling, Immunolabeling, TUNEL Assay, Microscopy

Journal: Archives of Medical Science : AMS

Article Title: The decrease in hippocampal transient receptor potential M2 (TRPM2) channel and muscarinic acetylcholine receptor 1 (CHRM1) is associated with memory loss in a surgical menopause rat model

doi: 10.5114/aoms.2019.83760

Figure Lengend Snippet: TRPM2 and CHRM1 gene expression levels were lower in G2 compared with G1 a Compared with G1 (p < 0.05).

Article Snippet: In order to prevent surface staining, after treatment with Ultra V Block (TA-125-UB, the Lab Vision Corporation, USA) solutions, the tissues were incubated with primary antibodies for 60 min (CHRM1 was purchased from Boster (Cholinergic receptor, muscarinic 1, catalog number: PA2202, Boster, 3942 B Valley Ave, Pleasanton, CA, 94566) and TRPM2 was purchased from Abcam (Rabbit Anti-TRPM2 antibody, ab101738, Abcam, Cambridge, UK)).

Techniques: Gene Expression

Journal: Archives of Medical Science : AMS

Article Title: The decrease in hippocampal transient receptor potential M2 (TRPM2) channel and muscarinic acetylcholine receptor 1 (CHRM1) is associated with memory loss in a surgical menopause rat model

doi: 10.5114/aoms.2019.83760

Figure Lengend Snippet: CHRM1, TRPM2 immunoreactivity and apoptotic histoscore (diffuseness × severity) and MDA levels

Article Snippet: In order to prevent surface staining, after treatment with Ultra V Block (TA-125-UB, the Lab Vision Corporation, USA) solutions, the tissues were incubated with primary antibodies for 60 min (CHRM1 was purchased from Boster (Cholinergic receptor, muscarinic 1, catalog number: PA2202, Boster, 3942 B Valley Ave, Pleasanton, CA, 94566) and TRPM2 was purchased from Abcam (Rabbit Anti-TRPM2 antibody, ab101738, Abcam, Cambridge, UK)).

Techniques: TUNEL Assay

Journal: Archives of Medical Science : AMS

Article Title: The decrease in hippocampal transient receptor potential M2 (TRPM2) channel and muscarinic acetylcholine receptor 1 (CHRM1) is associated with memory loss in a surgical menopause rat model

doi: 10.5114/aoms.2019.83760

Figure Lengend Snippet: A – Increased chrm1 immunoreactivity staining in hippocampal area is shown in G1 relative to G2 (black arrow). B – Increased trpm2 immunoreactivity staining in hippocampal area is shown in G1 relative to G2 (black arrow). C – TUNEL staining in hippocampal area is lower in G1 compared with G2 (black arrow). D – CHRM1 immunoreactivity staining in hippocampal area is lower in G2 compared with G1 (black arrow). E – TRPM2 immunoreactivity staining in hippocampal area is lower in G2 compared with G1 (black arrow). F – TUNEL staining in hippocampal area is higher in G2 compared with G1 (black arrow)

Article Snippet: In order to prevent surface staining, after treatment with Ultra V Block (TA-125-UB, the Lab Vision Corporation, USA) solutions, the tissues were incubated with primary antibodies for 60 min (CHRM1 was purchased from Boster (Cholinergic receptor, muscarinic 1, catalog number: PA2202, Boster, 3942 B Valley Ave, Pleasanton, CA, 94566) and TRPM2 was purchased from Abcam (Rabbit Anti-TRPM2 antibody, ab101738, Abcam, Cambridge, UK)).

Techniques: Staining, TUNEL Assay

Journal: BBA Clinical

Article Title: Novel panel of protein biomarkers to predict response to bortezomib-containing induction regimens in multiple myeloma patients

doi: 10.1016/j.bbacli.2017.05.003

Figure Lengend Snippet: Label-free mass spectrometry data. List of statistically significant discovered proteins using LC-MS analysis. Proteins (in bold) were selected for further validation using ELISAs.

Article Snippet: Four commercially available kits for these four proteins; angiogenin (ANG) [Abcam, UK - ab99970], clusterin (CLU) [R&D system, UK - DCLU00], C-C Motif Chemokine 18 (CCL18) [Abcam, UK - ab100620], and Complement C1q [Abcam, UK - ab170246] were used.

Techniques: Mass Spectrometry, Biomarker Discovery, Variant Assay

Journal: BBA Clinical

Article Title: Novel panel of protein biomarkers to predict response to bortezomib-containing induction regimens in multiple myeloma patients

doi: 10.1016/j.bbacli.2017.05.003

Figure Lengend Snippet: ELISA Data. Mean, SD, Area under the curve (AUC) and p -value for each of the new and standard proteins found in the two groups of patients compared.

Article Snippet: Four commercially available kits for these four proteins; angiogenin (ANG) [Abcam, UK - ab99970], clusterin (CLU) [R&D system, UK - DCLU00], C-C Motif Chemokine 18 (CCL18) [Abcam, UK - ab100620], and Complement C1q [Abcam, UK - ab170246] were used.

Techniques: Enzyme-linked Immunosorbent Assay

Journal: BBA Clinical

Article Title: Novel panel of protein biomarkers to predict response to bortezomib-containing induction regimens in multiple myeloma patients

doi: 10.1016/j.bbacli.2017.05.003

Figure Lengend Snippet: Logistic regression analysis data. List of different protein combinations used to establish the best model that can be used as a predictive panel for response to induction therapy containing bortezomib regime.

Article Snippet: Four commercially available kits for these four proteins; angiogenin (ANG) [Abcam, UK - ab99970], clusterin (CLU) [R&D system, UK - DCLU00], C-C Motif Chemokine 18 (CCL18) [Abcam, UK - ab100620], and Complement C1q [Abcam, UK - ab170246] were used.

Techniques: