Table 4. Data and statistical analysis from nuclear sensor replicates.
Top. Data represents biological replicates using transiently transfected nuclear sensor and nuclear cpVenus in HeLa cells. Data are arranged for input into STATA14 statistical software. “0” represents cells treated with 0 nM FK866. “1” represents cells treated with 10 nM FK866. The fl values represent the geometric mean from 488/405 nm fluorescence ratio of the healthy, single HeLa cells measured by cytometry. “experiment”, biological replicate; “fl”, 488/405 nm geometric mean fluorescent value; “treatment”, +/− 10 nM FK866; “sensor”, +/− nuclear NAD+ sensor.
Bottom. Calculated statistical interaction table from STATA14 using REML. The p-value for the ratio of ratios across replicates (bold red) is reported under the statistical interaction “_IsenXtre_1_1”. This represents the fluorescence change following FK866 treatment in the nuclear sensor, relative to changes in cpVenus. Calculating the exponential of the coefficient value, “Coef.”, represents the mean fold-change in fluorescence for the ratio of ratios across replicates. i.e. e^(0.42) = 1.52, 95% CI (e^0.25 – e^0.59) = 95% CI (1.28 to 1.80), p<0.001.
The fluorescence change following FK866 treatment in nuclear cpVenus across replicates is represented by “_Itreatment_1”: e^(coef) = e^(0.03) = 1.03, p=0.629.
To calculate the average fluorescent change in the nuclear sensor independent of cpVenus, combine the “treatment” and “sensor X treatment interaction” using the lincom command (step 50). The combined values (blue) represents the change in the sensor: e^(coef) = e^(0.45) = 1.57, 95% CI (1.39 to 1.77), p<0.001.
| experiment | fl | treatment | sensor |
|---|---|---|---|
| 1 | 9.97 | 0 | 0 |
| 1 | 10.00 | 1 | 0 |
| 1 | 3.78 | 0 | 1 |
| 1 | 6.45 | 1 | 1 |
| 2 | 6.83 | 0 | 0 |
| 2 | 6.85 | 1 | 0 |
| 2 | 2.26 | 0 | 1 |
| 2 | 3.34 | 1 | 1 |
| 3 | 6.70 | 0 | 0 |
| 3 | 7.28 | 1 | 0 |
| 3 | 2.30 | 0 | 1 |
| 3 | 3.56 | 1 | 1 |
| log_fl | Coef. | Std. Err. | z | P> |z| | [95% Conf. | Interval] |
|---|---|---|---|---|---|---|
| _Isensor_1 | −1.05 | 0.06 | −17.09 | 0 | −1.17 | −0.93 |
| _Itreatment_1 | 0.03 | 0.06 | 0.48 | 0.629 | −0.09 | 0.15 |
| _IsenXtre_1_1 | 0.42 | 0.09 | 4.89 | 0 | 0.25 | 0.59 |
| _cons | 2.04 | 0.16 | 12.73 | 0 | 1.73 | 2.36 |
| _IsenXtre_1_1 + _Itreatment_1 | 0.45 | 0.061337 | 7.40 | 0 | 0.33 | 0.57 |