Journal: BMC Neuroscience

Article Title: A role for prenylated rab acceptor 1 in vertebrate photoreceptor development

doi: 10.1186/1471-2202-13-152

Figure Lengend Snippet: Quantitative real-time PCR results for Pde6b and Rabac1 . The number of cycles required for detection of gene expression (right axis) of Pde6b ( A ) and Rabac1 ( B ) is indicated by blue diamonds (wt) and red triangles ( rd1 ) at each of the four time points between P2 – P8. For both genes, significantly more cycles were required for detection in the rd1 retinas at all time points, indicating that the genes are down-regulated. The yellow bars indicate the rd1 /wt ratio of expression (left axis). All data was normalized to the housekeeping gene HPRT.

Article Snippet: Membranes were then probed with a primary antibody to PRA1 (Proteintech Group, Chicago, IL; 1:500) overnight at 4°C.

Techniques: Real-time Polymerase Chain Reaction, Gene Expression, Expressing

Journal: BMC Neuroscience

Article Title: A role for prenylated rab acceptor 1 in vertebrate photoreceptor development

doi: 10.1186/1471-2202-13-152

Figure Lengend Snippet: Semiquantitative analysis of expression of PRA1 in wt and rd1 retina. Western blot analysis of PRA1 expression shows the protein is strongly downregulated in rd1 retinas at P2 an P4. Numbers below the blot represent the average relative fold difference of protein levels compared to P4 wt and normalized to β-actin on the basis of densitometer quantification of two independent experiments.

Article Snippet: Membranes were then probed with a primary antibody to PRA1 (Proteintech Group, Chicago, IL; 1:500) overnight at 4°C.

Techniques: Expressing, Western Blot

Journal: BMC Neuroscience

Article Title: A role for prenylated rab acceptor 1 in vertebrate photoreceptor development

doi: 10.1186/1471-2202-13-152

Figure Lengend Snippet: Localization of PRA1-LIR in wt and rd1 retina. Immunofluorescence labeling was performed on cryostat sections of mouse retinas harvested at P6 ( A – C ), P8 ( D – F ), P10 ( G – I ), and P21 ( J – L ). No staining was seen in control (ctl) sections processed without primary antibody ( A , D , G , and J ). In wt retinas ( B , E , H , and K ), PRA1-LIR was seen in photoreceptor inner segments (IS), particularly in the proximal region and in streaks through the outer nuclear layer (ONL) during differentiation, consistent with the growth of apical processes. These streaks were no longer present in the mature retina at P21 ( K ). In rd1 retinas ( C , F , I , and L ), PRA1-LIR was also present in IS but appeared less intense and more diffusely distributed. Insets, taken from an adjacent area to avoid bleaching, illustrate at 4× higher magnification the inner segment region, which is indicated between the bars on the right margin. In both genotypes, PRA1-LIR was seen in the perinuclear region of most inner retinal neurons and in both plexiform layers with increasing intensity during development. However, at P21 staining was more diffuse in the inner rd1 retina ( L ) compared to wt ( K ). Images are representative stacks of 6 confocal slices. Abbreviations: OPL = outer plexiform layer, INL = inner nuclear layer, IPL = inner plexiform layer, GCL = ganglion cell layer.

Article Snippet: Membranes were then probed with a primary antibody to PRA1 (Proteintech Group, Chicago, IL; 1:500) overnight at 4°C.

Techniques: Immunofluorescence, Labeling, Staining, Control

Journal: BMC Neuroscience

Article Title: A role for prenylated rab acceptor 1 in vertebrate photoreceptor development

doi: 10.1186/1471-2202-13-152

Figure Lengend Snippet: Relationship of PRA1-LIR to Golgi and cilia markers. ( A - C ) Immunofluorescent double labeling of PRA1-LIR and the cis-Golgi marker, GM-130-LIR, was performed on P10 mouse retinas. In control experiments ( A ), blood vessels stained non-specifically (arrowheads). In wt retinas ( B ), intense GM-130-LIR (magenta) co-localized with PRA1-LIR (green) in the proximal IS. Both overlapping and distinct punctae appeared within the streak-like pattern in the ONL (inset). Diffuse single staining of PRA1-LIR was seen in the distal IS. In rd1 retinas ( C ), GM-130-LIR was distributed more extensively in the IS (inset) with single (magenta arrowhead) and double (white arrowhead) labeled punctae in the distal IS. Images are single confocal slices. Insets illustrate the IS region, indicated between the white bars, at 4× higher magnification. ( D - F ) Immunofluorescent double labeling of Rab6-LIR (green) and GM-130-LIR (magenta) was performed on P10 wt ( D , E ) and rd1 ( F ) mouse retinas. Non-specific staining of blood vessels was present but not seen in the IS region illustrated ( D ). Rab6-LIR was closely apposed to GM-130-LIR with minimal overlap in both wt and rd1 . ( G , H ) Immunofluorescent double labeling of PRA1-LIR (green) and RP1-LIR (magenta) was performed on P21 wt mouse retinas. No staining was detected in control experiments ( G ). PRA1-LIR punctae were seen throughout the IS region and frequently in close proximity to RP1-LIR ( H , arrows), but not colocalizing. Very sparse PRA1 stained punctae were seen beyond RP1-LIR (arrowheads). Magnification for D - H is the same as insets in B and C . Abbreviations are the same as Figure .

Article Snippet: Membranes were then probed with a primary antibody to PRA1 (Proteintech Group, Chicago, IL; 1:500) overnight at 4°C.

Techniques: Labeling, Marker, Control, Staining

Journal: BMC Neuroscience

Article Title: A role for prenylated rab acceptor 1 in vertebrate photoreceptor development

doi: 10.1186/1471-2202-13-152

Figure Lengend Snippet: Quantitative analysis of PRA1-LIR intensity in wt and rd1 retinas. The intensity of the PRA1-like immunofluorescent signal in wt (blue) and rd1 (orange) was determined for the IS region ( A ) and the inner plexiform layer ( B ). PRA1-LIR was less intense in the IS layer of rd1 retinas when compared to age-matched wt retinas at all time points, although variability precluded significance at P6. In contrast, the average intensity of PRA1-LIR was very similar in wt and rd1 retinas with some difference seen only at P8. *p < 0.05, **p < 0.01.

Article Snippet: Membranes were then probed with a primary antibody to PRA1 (Proteintech Group, Chicago, IL; 1:500) overnight at 4°C.

Techniques:

Journal: bioRxiv

Article Title: AI-based antibody discovery platform identifies novel, diverse and pharmacologically active therapeutic antibodies against multiple SARS-CoV-2 strains

doi: 10.1101/2023.08.21.554197

Figure Lengend Snippet: Unpurified transfection supernatants were tested by AlphaLISA for binding to biotinylated SARS-CoV-2 spike protein (red symbols) and in parallel to biotinylated irrelevant target protein (black symbols). Positive control antibody M-3418 (S309) demonstrates binding specificity, whereas isotype control antibody demonstrates lack of binding. Replicates were included when possible. Data was graphed using GraphPad Prism software. Representative binding profile shown.

Article Snippet: Subsequently, 10 μl of Protein A AlphaLISA ® Acceptor beads (10 μg/ml final concentration; Perkin Elmer cat# AL101C) were added, and further incubated for 1 hour at room temperature in the dark with shaking (250 rpm).

Techniques: Transfection, Binding Assay, Positive Control, Control, Software

Journal: bioRxiv

Article Title: AI-based antibody discovery platform identifies novel, diverse and pharmacologically active therapeutic antibodies against multiple SARS-CoV-2 strains

doi: 10.1101/2023.08.21.554197

Figure Lengend Snippet: Binding screen allows identification of target-specific binders. Unpurified transfection supernatants were tested by AlphaLISA for binding to biotinylated SARS-CoV-2 Spike protein (red symbols) and in parallel to biotinylated irrelevant target protein (black symbols). Positive control antibody S309 demonstrates binding specificity, whereas isotype control antibody demonstrates lack of binding. to Replicates were included when possible. Data was graphed using GraphPad Prism software. Representative binding profiles are shown.

Article Snippet: Subsequently, 10 μl of Protein A AlphaLISA ® Acceptor beads (10 μg/ml final concentration; Perkin Elmer cat# AL101C) were added, and further incubated for 1 hour at room temperature in the dark with shaking (250 rpm).

Techniques: Binding Assay, Transfection, Positive Control, Control, Software

Journal: bioRxiv

Article Title: AI-based antibody discovery platform identifies novel, diverse and pharmacologically active therapeutic antibodies against multiple SARS-CoV-2 strains

doi: 10.1101/2023.08.21.554197

Figure Lengend Snippet: Unpurified transfection supernatants were concentration normalized and tested for binding in a 9-point serial titration series to biotinylated SARS-CoV-2 Spike protein by AlphaLISA. Replicates were included when possible. Data was graphed using GraphPad Prism software. Representative binding profiles for 30 J.HAL ® antibodies are shown in the sub-plots A-F above, along with the positive control antibody M-3418 (S309).

Article Snippet: Subsequently, 10 μl of Protein A AlphaLISA ® Acceptor beads (10 μg/ml final concentration; Perkin Elmer cat# AL101C) were added, and further incubated for 1 hour at room temperature in the dark with shaking (250 rpm).

Techniques: Transfection, Concentration Assay, Binding Assay, Titration, Software, Positive Control

Journal: bioRxiv

Article Title: AI-based antibody discovery platform identifies novel, diverse and pharmacologically active therapeutic antibodies against multiple SARS-CoV-2 strains

doi: 10.1101/2023.08.21.554197

Figure Lengend Snippet: Unpurified transfection supernatants were concentration normalized and tested for binding in a 9-point serial titration series to biotinylated SARS-CoV-2 Spike protein by AlphaLISA. Replicates were included when possible. Data was graphed using GraphPad Prism software. Representative binding profiles for 43 J.HAL ® antibodies are shown in the sub-plots A-F above, along with the positive control antibody M-3418 (S309).

Article Snippet: Subsequently, 10 μl of Protein A AlphaLISA ® Acceptor beads (10 μg/ml final concentration; Perkin Elmer cat# AL101C) were added, and further incubated for 1 hour at room temperature in the dark with shaking (250 rpm).

Techniques: Transfection, Concentration Assay, Binding Assay, Titration, Software, Positive Control