This repository contains the processed datasets and raw instrument exports supporting the manuscript “A Two-Point Fluorescence Detection Flow Cell with Empirical Calibration for High Optical Density Analysis” (manuscript under review; citation data will be updated pending publication). The study evaluates inner-filter-effect correction using two fixed fluorescence detection points (F₀, F₁) in a custom flow cell under (i) static sample loading and (ii) continuous-flow gradient conditions, with additional datasets probing flow-rate dependence and fluorophore-specific behaviour. Contents 1) Processed datasets (Excel) The Excel workbooks contain processed fluorescence data, calibration workflows, and reported performance metrics (R², LOD, LOD%, mErr%). Weitner and Sakic – Fluorescence Flow Cell – FL-STATIC.xlsxStatic (manual) sample loading dataset for fluorescein, including concentration series, extracted fluorescence signals (F₀, F₁), corrected signal (FIFE_CORR), and calibration metrics used in the main manuscript. Weitner and Sakic – Fluorescence Flow Cell – FL-FLOW-1.0mLmin-1.xlsxContinuous-flow gradient dataset for fluorescein at 1.0 mL min⁻¹, including UV-corrected concentration axis, extracted fluorescence signals, corrected signal, and calibration metrics corresponding to the main manuscript figures. Weitner and Sakic – Fluorescence Flow Cell – FL-FLOW-0.5mLmin-1.xlsxAdditional fluorescein dataset at reduced flow rate (0.5 mL min⁻¹), included to assess the influence of hydrodynamic conditions on calibration parameters and performance. Weitner and Sakic – Fluorescence Flow Cell – QS-FLOW-0.5mLmin-1.xlsxQuinine sulfate dataset (0.5 mL min⁻¹), included to evaluate the applicability of the method to fluorophores with different spectral properties and lower signal-to-noise ratio. 2) Electronic Supplementary Information (ESI) ESI document (PDF)Provides extended methodological and computational details, including: flow-cell fabrication and optical considerations effective optical path length calibration model and Excel Solver implementation dataset structure and processing workflow UV-based gradient reconstruction and delay correction additional datasets (flow-rate variation and quinine sulfate) These materials enable full reproducibility of the calibration procedure and interpretation of the datasets. 3) Raw instrument exports (ASCII)These files are provided to enable full reprocessing from primary instrument outputs. (a) ÄKTA start exports (gradient method + run log) Method export (UNICORN start 1.1): tabulated parameter list for the programmed gradient, including flow rate, mixer/delay volume parameters, and gradient segment definitions (0–100% B). Raw run data export: time/volume series, including UV (mAU), gradient concentration (%B), system flow, pressure, and run log annotations. (b) Ocean Optics / Ocean Insight exports (spectral time series) ASCII spectral time-series exports recorded during gradient runs, including acquisition metadata (spectrometer ID, trigger mode, scans-to-average, integration time, pixel count) followed by wavelength-resolved spectra. These files provide the raw fluorescence spectra from which F₀/F₁ intensities were extracted. Notes on gradient verification and correction The effective gradient delivered to the flow cell was verified using on-line UV absorbance at 280 nm (ÄKTA UV detector, 0.2 cm path length). Due to non-linearity at the gradient endpoints (valve duty-cycle limitations and mixer dispersion), only the central region was used for IFE analysis. The measured UV trace was used to construct a corrected concentration axis by mapping the recorded signal onto an ideal linear gradient. Reuse The Excel workbooks allow direct reproduction of the manuscript figures and calibration results. In this updated release, all datasets have been realigned and reprocessed using the Excel Solver add-in. This includes reconstruction of the effective concentration axis (for flow datasets), recalculation of corrected fluorescence signals, and re-optimization of calibration parameters.