Journal:

Article Title: Studies of a germinal centre B-cell expressed gene, GCET2 , suggest its role as a membrane associated adapter protein

doi: 10.1111/j.1365-2141.2007.06597.x

Figure Lengend Snippet: Localisation of germinal centre B-cell expressed transcript 2 (GCET2) in COS7 and B cells by confocal microscopy. (A) Localisation of GCET2 in COS7 cells. This figure shows the negative control. (B) Localisation of GCET2 in COS7 cells by confocal analysis. Wild-type GCET2 was mainly localised in the cell membrane with some distribution in the cytosol, probably in the Golgi apparatus and the membrane of cytosolic organelles. (C) Localisation of wild-type GCET2 in COS7 cells. (D) Localisation of GCET2 in DHL16 cells. DHL16 cells without pMIG-GCET2 transduction were used as negative control. Cells were stained using mouse anti-V5 monoclonal primary antibody and Texas red conjugated horse anti-mouse IgG secondary antibody. This picture is the Differential Interface Contrast (DIC) of control DHL16 B cells. (E) Fluorescence of control DHL16 cells. The cells were totally negative for red signals. (F) Combination (overlay) of G and H. (G) DIC of pMIG-GCET2 transduced B cell. (H) Red fluorescence of pMIG-GCET2 transduced B cells. Red signals were predominantly in the cell membrane and the Golgi area. (I) Combination (overlay) of J and K.

Article Snippet: Slides were incubated in anti-V5 primary antibody (5 µg/ml) for 1 h at room temperature with gentle agitation, and in Texas Red conjugated horse anti-mouse IgG secondary antibody (7·5 µg/ml; Vector Laboratories) for 1 h at room temperature in the dark with gentle agitation.

Techniques: Confocal Microscopy, Negative Control, Transduction, Staining, Fluorescence

Journal:

Article Title: Chlamydia trachomatis infection modulates trophoblast cytokine/chemokine production 1

doi: 10.4049/jimmunol.0800764

Figure Lengend Snippet: Inclusion formation and infection rates in trophoblast cells infected with C. trachomatis. (A) Trophoblast cells (Sw.71) were exposed to either No infection (NI), Ct serovar D (Ct-D) or Ct serovar L1 (Ct-L1) at a MOI of 1, by rocking/resting at room temperature for 2 hour. Inclusion formation was evaluated by light microscopy at 24 and 36 hours post-infection for Ct-L1 and Ct-D, respectively. Inclusions are highlighted by arrow heads (Mag. ×40). (B) After 36 hours of infection with or without Ct-L1 or Ct-D, inclusion formation in the Sw.71 and H8 cells was evaluated by by staining cells intracellularly with a FITC-conjugated mouse anti-Ct LPS mAb. Infection rates were then determined by both immmunofluorescent microscopy and flow cytometry. The flow cytometry histograms show two distinct populations: the left hand peak being the uninfected cells, and the right hand peak with the marker representing the infected trophoblast population.

Article Snippet: Following this incubation, membranes were washed three times as before and then incubated at room temperature for 1 hour with the horse anti-mouse secondary antibody conjugated to peroxidase (Vector Labs) in PBS-T/1% FFPM.

Techniques: Infection, Light Microscopy, Staining, Microscopy, Flow Cytometry, Marker

Journal:

Article Title: Chlamydia trachomatis infection modulates trophoblast cytokine/chemokine production 1

doi: 10.4049/jimmunol.0800764

Figure Lengend Snippet: Infection rates of trophoblast cells infected with C. trachomatis by different techniques. (A) H8 and Sw.71 cells were infected with Ct (serovar D) at an MOI of 1 by either rocking or by centrifugation. After 36 hours, the cells were collected and stained intracellularly with a FITC-conjugated mouse anti-Ct LPS mAb. Centrifugation resulted in a higher rate of infection in both cell lines (B) H8 and Sw.71 cells were infected with or without Ct (serovar D) at an MOI of 1 by centrifugation. After 48 and 72 hours inclusion formation was visualized by light microscopy. Inclusions contained within the cells are highlighted by arrow heads, while extruded inclusions are highlighted by asterisks (Mag. ×40).

Article Snippet: Following this incubation, membranes were washed three times as before and then incubated at room temperature for 1 hour with the horse anti-mouse secondary antibody conjugated to peroxidase (Vector Labs) in PBS-T/1% FFPM.

Techniques: Infection, Centrifugation, Staining, Light Microscopy

Journal:

Article Title: Chlamydia trachomatis infection modulates trophoblast cytokine/chemokine production 1

doi: 10.4049/jimmunol.0800764

Figure Lengend Snippet: Chlamydia-infected trophoblast cells produce viable EBs. H8 and Sw.71 cells were infected with Ct (serovar D) at an MOI of 1 by centrifugation. After 36 hours, the cells were collected and stained intracellularly with a FITC-conjugated mouse anti-Ct LPS mAb. The infection levels were then determined by flow cytometry (i & iv). In parallel, lysates were prepared from Ct-infected H8 and Sw.71 cells and these were then immediately applied to a culture of uninfected HeLa cells. After 48 hours, the HeLa cells exposed to infected Sw.71 or H8 lysates were collected and the infection levels determined by flow cytometry. Histograms (ii & v) show the levels of HeLa cell Ct infection (solid line), when compared to the uninfected HeLa cells (dotted line). The HeLa cells exposed to infected Sw.71 or H8 lysates were also evaluated for inclusion formation by light microcopy (iii & vi). Inclusions contained within the cells are highlighted by arrow heads, while extruded inclusions are highlighted by asterisks (Mag. ×40) (iii & iv).

Article Snippet: Following this incubation, membranes were washed three times as before and then incubated at room temperature for 1 hour with the horse anti-mouse secondary antibody conjugated to peroxidase (Vector Labs) in PBS-T/1% FFPM.

Techniques: Infection, Centrifugation, Staining, Flow Cytometry