Relative light units

Coelenterazine Analogue used Initial Rate of Regeneration (in relative light units/s)... 

The normalized luciferase activity is calculated by dividing the relative light units (RLU) of firefly luciferase activity-background luminescence by that of Renilla luciferase-background luminescence. Typically, the cell lysate background is around 200 to 300 RLU (similar to that of a buffer control) and about 104 to 105 and 5 to 20 x 103 RLU for firefly and Renilla luciferase luminescence, respectively. 

Relative Light Units / jug 31 Optical Density of DNA in Solution (260 nm)... 

Figure 6 Lipofection results (lipofection profiles) of lipoplexes from the R-configu-rated cationic lipids KL-1-1 to KL-1-17 (Table 1) in a mixture with equimolar amounts of l,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine (DOPE) (counterion chloride) and the pCMVluc-plasmid. Each bar represents the mean ( S.D.) of three wells of a 96-well microtiter plate. T-axis (left) represents the transfection efficiencies expressed in relative light units (RLU) (lu/pg protein). X-axis (right) represents the viability of the cells compared to nontreated control cells. F-axis represents the different cationic lipid/plasmid DNA-charge ratios from 1 to 15.

Abbreviations DOPE, l,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine DOTAP, iV-[l-(2, 3-dioleoyloxy)propyl]-A,iV,JV-trrmethylanimoniumchloride TE, transfection efficiency DC, dendritic cells RLU, relative light units chol, cholesterol. 

On each 96-well microtiter plate, a eomplete 2,3,7,8-TCDD standard concentration range was incubated and analyzed in triplieate. A eurve fit of the 2,3,7,8-TCDD standard range was produced for the calculation of DR CALUX TEQ content in the samples tested. The analyzed relative light units (RLU) from the samples were interpolated on the 2,3,7,8-TCDD standard curve, and the DR CALUX TEQ content was quantified between the limit of quantitation (LOQ) and the concentration of 2,3,7,8-TCDD at whieh 50% of the maximum response is observed (EC50). 

The relative light units have been corrected for dimethyl sulfoxide blank. r O.993. 

Figure 16.6 Polyfection mediated by a disulfide-containing poly lysine conjugate. (A) 293-T7 cells and (B) HepG2 cells were incubated for four hours at 37 °C with pCMVluc (5 pig) complexed with either disulfide-containing polylysine (20 pig) or polylysine (15 pig) in the presence of 10% FCS and 100 iM chloroquine. The luciferase activity was measured after 48 hours culture and expressed as the relative light units (RLU) per 106 cells.

The luminescence data, normalized over protein content of each sample, are expressed as relative light units (RLU) per xg of protein. 

Spin down cells and remove media. Lyse cells and perform reporter assay as in Subheading 3.1.3. to assay for relative light units as readout for luciferase activity. 

Incubate cells with transfection complex for 24 h. Test cell viability using 0.4% Trypan Blue to stain cells and count viable cells at seeding, day of transfection, and just prior to harvest. Harvest cells and assay for reporter gene activity Spin down cells and lyse cells to assay for luciferase activity. Figures 1 and 2 illustrate typical transfection results, observed visually as lacZ stained cells (Fig. 1) or Relative Light Units (RLUs) when luciferase activity is measured (Fig. 2). 

Fig. 2. TransIT-LTl-mediated transfection of HeLa cells at varying cell densities HeLa cells were plated in 12-well plates and transfected in parallel at 50% and 90% confluency. TransIT-LTl reagent transfections were performed in duplicate using a luciferase expression vector (pCI-luc) and 3 iL reagent per well, shows the importance of plating cells at optimal density for transfection. Twenty-four hours post-transfection, cells were harvested and assayed for luciferase activity. Visual confluence (line graph) was measured under the microscope at harvest. The data represent the average luciferase activity (Relative Light Units - RLUs in Millions) from three experiments performed on different days.

Assay using relative light units produced by luciferase as the read out by the method described for DNA transfection reporter assay in Subheading 3.3.5. Other methods of choice to determine gene knockdown are Northern blots, RT-PCR, RNase protection, and branched DNA assays, apart from which assays to measure target protein produced can also be used. 

Fig. 12 Transfection activities of the DMAEC-SS-polyrotaxanes, DMAEC-polyrotaxanes, and the LPEI22k at different N/P ratios in NIH/3T3 cells. Luciferase activity in the NIH/3T3 cells was measured at 48 h after the addition of the polyplexes. Results were expressed as relative lights units (RLU) per mg of cell protein...

Luciferase activity in the supernatant was measured using a luciferase assay system and a luminometer. The activity is reported in relative light units (RLU) per mg protein of cells or tissue. 

Functional characterization of the new photoprotein was carried out by biochemical analysis of purified recombinant protein. Different quantities of recombinant Photina ranging from 0.09 to 12.5 ng were charged with 10 0M coelenterazine for 4 h at 4 °C. After incubation, 100 pM CaCl2 was applied and the total RLU (relative light units) was recorded for 10 s using a Berthold Luminometer. As shown in Fig. 3, recombinant Photina gives a consistent signal even when used at very low concentrations. 

Fig. 2. (A) Effects of mepyramine on the basal reporter-gene (luciferase) activity in mock transfected COS-7 cells (O) or in COS-7 cells transiently expressing the human histamine H j receptor at 1 0.1 ( ), 2.8 0.1 (O) and 4.2+ 0.2 ( ) pmol/mg protein. (B) Effects of mepyramine on the basal luciferase activity in COS-7 cells transiently transfected with the human histamine H, receptor in the absence ( ) or presence of either 1.0 p.g (O), 5.0 p,g ( ), or 25 xg ( ) pCMVGaii /lO cells. RLU = relative light units.

Fig. 8.2 Transfection activities of multifunctional envelope-type nano devices (MENDs). NIH3T3 cells were transfected with different particles and MENDs containing a luciferase coding plasmid DNA. Luciferase activities were expressed as the relative light unit (RLU) per mg of protein.

FIGURE 6.8 Transfection efficiency (quantified as relative light units [RLU]/mg protein) of six different SLN formulations (Co DDAB 4% Compritol ATO 888, 1% dimethyldiocta-decylammonium bromide = DDAB Co EQ 4% Compritol, 1% N,N-di-(P-stearoylethyl)-N,N-dimethyl-ammonium chloride = EQ Co DOTAP 4% Compritol, 1% N-[l-(2,3-dioleoy-loxy)propyl]-N,N,N-trimethylammonium chloride = DOTAP Cp DDAB 4% cetyl pahnitate, 1% DDAB Cp EQ 4% cetyl pahnitate, 1% EQ Cp DOTAP 4% cetyl palmitate, 1% DOTAP) all formulations were stabilized with 2% [7 3] Tween 80 Span 85). The experiments were performed on Cos-1 cells (African green monkey kidney). The transfection efficiency of naked plasmid (DNA alone) served as control. All tested SLN-DNA complexes showed a statistically significant increase of transfection activity compared to naked plasmid. Cp DOTAP was 10 times more potent then the other formulations. 

FIGURE 6.19 Transfection efficiency (quantified as relative light units [RLU]/mg protein) of three different SLN formulations (all made from 4% cetyl pahnitate, 2% Tween 80/Span 85 [7 3], and 0.5% [S0.5], and 1.0% [SI] or 2.0% [S2] of the cationic hpid DOTAP) without medium (Medium) and with medium and 100 piM chloroquine phosphate (Med. + 100 J,M QC). The chloroquine addition enhanced transfection activity for all tested SLN in different extends. 

Light emitted by chemiluminescent substrate was expressed in relative light units (RLU). The time dependence of light intensity was measured and the peak intensity was converted into percent of inhibition relative to control. We expressed the results as total antioxidant potential, expressed as percent inhibition TAPcl = ((max RLUconiroi) - (max RLUsampie)) 100 / (max RLUcontroi). Figure 1 shows the correlation between sum of phenolic compounds... 

Figure 3.19 Delivery of plasmid DNA by f-SWNTs and expression in cells. Levels of marker gene (b-gal) expression in CHO cells in relative light units (RLU) per mg total protein. Reprinted with permission from ref. 170. Copyright 2004 Wiley-VCH Verlag GmbH Co. KGaA.

Interpretations of the assay are based on the ratio of the response (in relative light units) of the specimen and the mean of three negative controls. The Leader luminometer calculates the results automatically and prints a hard-copy result for each test sample. This Gen-Probe chemiluminescence assay may be used to detect the presence of N. gonorrhoeae directly in urogenital samples or to confirm the identity of possible gonococcal isolates. 

The results of the Gen-Probe PACE 2 system for Neisseria gonorrhoeae are calculated based on the difference between the response in relative light units (RLU) of the specimen and the mean of the negative reference. 

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