One published dataset, run through Ampliova.
The raw RT-qPCR files behind Figure 2A of O'Callaghan et al. (2026), taken from figshare and loaded into Ampliova unchanged. Human iNeurons from four inductions were grown in two culture media, and PINK1 was measured against two reference genes. Click a bar to see its wells, then any well to open its curve.
Data and licence
O'Callaghan et al. (2026). Glucose concentration of neuronal media formulations influences PINK1-dependent mitophagy in human iNeurons.
Raw data on figshare (doi:10.5522/04/29064815), licence CC BY 4.0. Preprint doi:10.1101/2025.04.01.646648.
How the plates were set up
Every Cq and every curve comes straight from the files. The wells were labelled the way their names describe them. UBC and RPL18A are the reference genes, wells named NTC_ or NRT_ plus a sample name are no-RT controls, and NTC_water wells are no-template controls. Plate 3 has one oddity. Well H11 carries a sample name but sits in the water row next to the other no-template controls, so it was treated as one.
Against the published figure
The authors normalised to UBC alone with 2^-ΔΔCq and saw no difference in PINK1 mRNA between the two media (n = 4 inductions, t-test). Ampliova corrects each gene for its own efficiency and normalises to both reference genes. That puts BrainPhys at 1.15× N2B27 (ratio of geometric means, 95% CI 0.98 to 1.36; Welch's t-test on log2, P = 0.0736). Like the authors, Ampliova finds no significant difference. The interval still runs from a 2% decrease to a 36% increase, so the data do not show that the two media have the same effect either.
| Sample | Group | Fold change | Norm. qty | |
|---|---|---|---|---|
| 19_IND4_D24_N2B27 | N2B27 | 0.93× | 0.02728 | |
| 29_IND5_D24_N2B27 | N2B27 | 1.00× | 0.02950 | |
| 9_IND3_D24_N2B27 | N2B27 | 1.16× | 0.03402 | |
| N4_N2B27 | N2B27 | 0.91× | 0.02670 | |
| 10_IND3_D24_BrainPhys | BrainPhys | 1.23× | 0.03601 | |
| 20_IND4_D24_BrainPhys | BrainPhys | 1.13× | 0.03314 | |
| 30_IND5_D24_BrainPhys | BrainPhys | 1.15× | 0.03386 | |
| N4_BrainPhys | BrainPhys | 1.09× | 0.03205 | |
| Mean BrainPhys | 1.15× | |||
| Gene | geNorm M | BestKeeper MAD | BestKeeper CV (%) | ΔCq SD | NormFinder | Combined rank |
|---|---|---|---|---|---|---|
| RPL18A★ Most stable | 0.622#1 | 0.677#2 | 3.9 | 0.622#1 | 0.311 ⚠#1 | 1 |
| UBC | 0.622#2 | 0.424#1 | 2.2 | 0.622#2 | 0.311 ⚠#2 | 2 |
From a bar to its wells
Click a bar, or pick a group below. Under each sample are the wells that went into it, and every well opens its amplification curve.
Pick a group to list its samples and wells.
Efficiency, well by well
LinRegPCR fits a curve to each well. After an iterative Grubbs test drops the outlying wells, the mean of what is left becomes the gene's efficiency in the calculation.
| Gene | Wells | Range | Mean used | Excluded by Grubbs |
|---|---|---|---|---|
| PINK1 | 16 | 1.783 to 1.974 | 1.827 | plate 3 · D7 · 1.974 |
| UBC | 16 | 1.754 to 2.117 | 1.826 | plate 2 · C5 · 2.117 |
| RPL18A | 16 | 1.532 to 1.883 | 1.739 | none |
All 48 well efficiencies
PINK1
P1 C71.850
P1 C81.786
P1 D71.798
P1 D81.789
P2 C71.830
P2 C81.828
P2 D71.818
P2 D81.828
P3 C71.808
P3 C81.828
P3 D71.974
P3 D81.783
P4 C71.862
P4 C81.859
P4 D71.848
P4 D81.883
UBC
P1 C41.796
P1 C51.895
P1 D41.805
P1 D51.895
P2 C41.774
P2 C52.117
P2 D41.788
P2 D51.770
P3 C41.818
P3 C51.754
P3 D41.954
P3 D51.815
P4 C41.788
P4 C51.779
P4 D41.877
P4 D51.879
RPL18A
P1 C11.532
P1 C21.563
P1 D11.844
P1 D21.562
P2 C11.801
P2 C21.742
P2 D11.667
P2 D21.854
P3 C11.883
P3 C21.834
P3 D11.786
P3 D21.823
P4 C11.814
P4 C21.686
P4 D11.803
P4 D21.627
Methods paragraph
Drafted from the batch settings, the same text a Researcher-plan user gets. In your own batches it is the data-analysis section of an editable Methods tab, next to your bench protocol.
qPCR data were analyzed using Ampliova (ampliova.com). The target gene analyzed was PINK1. Per-reaction amplification efficiencies were derived from raw amplification curves using LinRegPCR (Ruijter et al., 2009, Nucleic Acids Res; Untergasser et al., 2021, BMC Bioinformatics), following the efficiency-corrected relative quantification protocol of Müller and Tiret (2026, Bio-Protocol). Efficiency outliers were identified among the per-well efficiencies of each gene and excluded using an iterative two-sided Grubbs test (P < 0.05), applied only to genes with at least six usable wells, and the mean efficiency of the remaining wells was used for each gene. Mean amplification efficiencies (E) were: UBC = 1.826, PINK1 = 1.827, RPL18A = 1.739. Expression levels were normalized to the geometric mean of 2 reference genes (UBC and RPL18A) (Vandesompele et al., 2002, Genome Biology). Fold changes were calculated relative to the N2B27 group. Analyses were performed across 4 plates, without inter-run calibration. Statistical analysis used one value per biological sample, technical replicates having been averaged on the Cq scale, and was performed on log2-transformed fold changes, on which the multiplicative measurement error of qPCR is approximately additive and homoscedastic. All tests were two-sided with α = 0.05. PINK1 expression was compared between N2B27 (n = 4) and BrainPhys (n = 4) using Welch's t-test. Effects are reported as ratios of geometric means with 95% confidence intervals. The test was specified in advance rather than selected from assumption checks; normality of the residuals (Shapiro-Wilk) and, between independent groups, equality of variances (Brown-Forsythe) were examined and reported. Statistics were computed with SciPy 1.14.1 and NumPy 1.26.4 (Virtanen et al., 2020, Nat Methods). Efficiency correction and normalization to multiple reference genes follow the MIQE 2.0 recommendations (Bustin et al., 2025, Clin Chem).
Full report
The MIQE 2.0 checklist, tables and figures, generated from this batch by the same code as the export button.