mouse anti-ccl2 antibody (clone 4b8)  (Probiodrug)

Journal: Acta Neuropathologica

Article Title: Isoglutaminyl cyclase contributes to CCL2-driven neuroinflammation in Alzheimer’s disease

doi: 10.1007/s00401-015-1395-2

Figure Lengend Snippet: Cocktails of antibodies used for double and triple labeling immunohistochemistry

Article Snippet: The goat anti-CCL2 antiserum sc-1784 (St. Cruz) and the mouse anti-CCL2 antibody (clone 4B8; Probiodrug, Halle/Saale, Germany) showed a distinct staining pattern for CCL2 around nuclei of mouse primary neurons as well as of neurons in mouse brain tissue.

Techniques: Labeling

Journal: Acta Neuropathologica

Article Title: Isoglutaminyl cyclase contributes to CCL2-driven neuroinflammation in Alzheimer’s disease

doi: 10.1007/s00401-015-1395-2

Figure Lengend Snippet: Neuronal isoQC expression, subcellular localization and co-expression of CCL2 in mouse brain and primary neurons. a IsoQC was strictly co-localized with the neuronal marker HuC/D in brain sections of 17-month-old wild-type mice demonstrating a neuron-specific expression. Cells which did not display HuC/D immunoreactivity were also negative for isoQC labeling ( arrows ). b IsoQC was co-localized with cellular compartment markers calreticulin and syntaxin-6, consistent with a subcellular localization in endoplasmic reticulum and Golgi apparatus. c The putative isoQC substrate CCL2 was found to be co-expressed by isoQC-immunoreactive neurons in cortex of wild-type mice ( top ) and in primary neuronal cultures ( bottom ). d The conversion of CCL2 by recombinant isoQC was analyzed in a kinetic assay. The progress curve ( black trace ) was in accordance with a curve modeled according to the integrated form of the Michaelis–Menten equation ( red trace ), enabling the determination of the kinetic parameters K M (19.8 ± 0.4 μM) and k cat (0.76 ± 0.01 s −1 )

Article Snippet: The goat anti-CCL2 antiserum sc-1784 (St. Cruz) and the mouse anti-CCL2 antibody (clone 4B8; Probiodrug, Halle/Saale, Germany) showed a distinct staining pattern for CCL2 around nuclei of mouse primary neurons as well as of neurons in mouse brain tissue.

Techniques: Expressing, Marker, Labeling, Recombinant, Kinetic Assay

Journal: Acta Neuropathologica

Article Title: Isoglutaminyl cyclase contributes to CCL2-driven neuroinflammation in Alzheimer’s disease

doi: 10.1007/s00401-015-1395-2

Figure Lengend Snippet: Regulation of isoQC and CCL2 expression in Tg2576 mice. a The neocortical isoQC and CCL2 mRNA levels were increased by 135 and 55 %, respectively, in 17-month-old APP transgenic Tg2576 mice ( black bars ) compared to wild-type littermates ( white bars ) as demonstrated by qRT-PCR analyses. Additionally, GFAP mRNA levels were increased by 115 %, indicating astrogliosis in Tg2576 mice. The isoQC and pGlu-CCL2 protein levels, however, were not affected in Tg2576 mice as measured by Western blot analysis and ELISA, respectively. b In 17-month-old wild-type (wt) and APP transgenic Tg2576 mice (tg) there was a predominantly neuronal expression of isoQC and CCL2 ( green immunofluorescence) as revealed by co-expression of HuC/D ( red immunofluorescence). The asterisks indicate the position of Abeta plaques in Tg2576 tissue. c In addition, aged Tg2576 mice—but not wild-type littermates—displayed astrocytic expression of isoQC and CCL2 in proximity of Abeta deposits following a gradient from the core towards the periphery of plaques ( I , within plaque core diameter; II , double plaque core diameter; III , triple plaque core diameter). d The astrocytic co-expression ( arrows ) of isoQC and CCL2 in proximity of Abeta plaques ( asterisks ) is visualized by the co-expression of the astrocyte marker GFAP ( red immunofluorescence). *p < 0.05, **p < 0.01, ***p < 0.001

Article Snippet: The goat anti-CCL2 antiserum sc-1784 (St. Cruz) and the mouse anti-CCL2 antibody (clone 4B8; Probiodrug, Halle/Saale, Germany) showed a distinct staining pattern for CCL2 around nuclei of mouse primary neurons as well as of neurons in mouse brain tissue.

Techniques: Expressing, Transgenic Assay, Quantitative RT-PCR, Western Blot, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Marker

Journal: Acta Neuropathologica

Article Title: Isoglutaminyl cyclase contributes to CCL2-driven neuroinflammation in Alzheimer’s disease

doi: 10.1007/s00401-015-1395-2

Figure Lengend Snippet: Co-regulation of isoQC and CCL2 in mouse primary astrocytes upon Abeta stimulation. a Immunocytochemical double labeling of isoQC ( green ) and CCL2 ( red ) with nuclear Hoechst counterstaining ( blue ) under control conditions and after stimulation with Abeta (5 μM) or pGlu-Abeta (5 μM) as indicated. Note the robust increase in the immunocytochemical labeling intensity for both proteins at 24 and 48 h and the decline at 72 h. b Quantification of immunocytochemical labeling revealed a highly significant time- and Abeta peptide-specific increase in isoQC and CCL2 immunoreactivity. Correlation analyses between isoQC and CCL2 immunocytochemical labeling demonstrated highly significant correlations in the expression of enzyme and substrate under control conditions ( b1 ), after stimulation with Abeta ( b2 ) and with pGlu-Abeta ( b3 ). c Quantification of isoQC and CCL2 mRNA expression under control conditions and after stimulation with Abeta and pGlu-Abeta for different periods of time as indicated ( N = 6 per time point). Note the absence of an increase of isoQC or CCL2 mRNA transcripts independent of the type and duration of the treatment. Correlation analyses of CCL2 mRNA levels plotted versus isoQC mRNA levels in individual astrocyte culture wells under control conditions and after (pGlu)-Abeta stimulation including all time points analyzed. Note the absence of a correlation between enzyme and substrate mRNA expression under control ( c1 ) and treatment conditions ( c2 , c3 )

Article Snippet: The goat anti-CCL2 antiserum sc-1784 (St. Cruz) and the mouse anti-CCL2 antibody (clone 4B8; Probiodrug, Halle/Saale, Germany) showed a distinct staining pattern for CCL2 around nuclei of mouse primary neurons as well as of neurons in mouse brain tissue.

Techniques: Labeling, Control, Expressing

Journal: Acta Neuropathologica

Article Title: Isoglutaminyl cyclase contributes to CCL2-driven neuroinflammation in Alzheimer’s disease

doi: 10.1007/s00401-015-1395-2

Figure Lengend Snippet: Immunohistological alterations of isoQC and CCL2 expression in AD cortex. a Immunohistochemistry for isoQC in human temporal cortex revealed a weak neuronal expression in control subjects. In corresponding tissue sections of AD subjects, an increased neuronal labeling intensity, in particular in layer III pyramidal neurons was observed. b A similar up-regulation in pyramidal layer III neurons was observed for CCL2. Additionally, both isoQC and CCL2 were induced in glia-like cells in proximity of Abeta plaques ( insets in a and b ). c Triple immunofluorescent labelings of isoQC with CCL2 and GFAP identified these glial cells as astrocytes surrounding Abeta deposits. d The astrocytic expression of isoQC and CCL2 in proximity of Abeta deposits followed a gradient from the core to the periphery of plaques ( I , within plaque core diameter; II , double plaque core diameter; III , triple plaque core diameter). **p < 0.01, ***p < 0.001

Article Snippet: The goat anti-CCL2 antiserum sc-1784 (St. Cruz) and the mouse anti-CCL2 antibody (clone 4B8; Probiodrug, Halle/Saale, Germany) showed a distinct staining pattern for CCL2 around nuclei of mouse primary neurons as well as of neurons in mouse brain tissue.

Techniques: Expressing, Immunohistochemistry, Control, Labeling

Journal: Acta Neuropathologica

Article Title: Isoglutaminyl cyclase contributes to CCL2-driven neuroinflammation in Alzheimer’s disease

doi: 10.1007/s00401-015-1395-2

Figure Lengend Snippet: Characteristics of isoQC, CCL2 and pGlu-Abeta accumulation in AD and correlation analyses with MMSE. a Quantitative analyses revealed statistically significant increases of isoQC and CCL2 mRNA levels as well as pGlu-CCL2 protein in temporal cortex samples from AD cases compared to control subjects ( *p < 0.05). b There was a statistically significant correlation between higher isoQC mRNA levels and the decline in MMSE ( r = −0.7220; p = 0.0080). c Likewise, a statistically significant correlation between higher CCL2 mRNA levels and the decline in MMSE ( r = −0.6124; p = 0.0343) was detected. d There was no correlation between isoQC and CCL2 transcript levels in the human brain samples analyzed. e A strong positive correlation between CCL2 mRNA and protein levels was detected ( r = 0.8552; p = 0.0004). f As for CCL2 mRNA, an inverse correlation between pGlu-CCL2 protein and MMSE was revealed ( r = −0.6336; p = 0.0270). While there was no correlation between isoQC mRNA levels and total Abeta42 peptide concentrations ( g ), a significant correlation between isoQC transcript levels and pGlu-Abeta concentrations was established ( r = 0.6663; p = 0.0180) ( h )

Article Snippet: The goat anti-CCL2 antiserum sc-1784 (St. Cruz) and the mouse anti-CCL2 antibody (clone 4B8; Probiodrug, Halle/Saale, Germany) showed a distinct staining pattern for CCL2 around nuclei of mouse primary neurons as well as of neurons in mouse brain tissue.

Techniques: Control