EV Phenotyping Interpretation Tool

This EV-iTEC tool supports the interpretation of extracellular vesicle (EV) phenotyping data obtained by NanoFCM, CytoFLEX nano and related single-particle flow cytometry platforms.

The tool provides background-corrected marker positivity estimates, confidence intervals, marker-profile summaries and QC-oriented interpretation guidance for common EV-associated markers.

Scientific disclaimer

This tool is intended for scientific and educational use only. Marker positivity, ratios, confidence intervals and Poisson-derived estimates are operational approximations and should not be interpreted as absolute EV identity, antigen copy number or biological function. Thresholds and interpretation rules represent empirical EV-iTEC guidance values and are not validated consensus cut-offs. Results should always be interpreted together with appropriate controls, orthogonal validation methods and biological context.

Input

All values should refer to detected events within the same EV gate and marker-positive gate.

Experiment

Marker

Instrument

Detected events

Total detected EV events after trigger/EV gate

Marker-positive events events above the marker gate

Background-positive events unstained / isotype / antibody-only in the same marker gate

Background total events total events in the background control

Concentration / acquisition

Total particle concentration particles/ml, optional; ideally same instrument/gate

Observed event rate events/s, optional

Event separation assumption advanced; ms; leave default if unknown

Dilution linearity checked?

Optional control

Detergent sensitivity % event loss after Triton/NP-40; optional

Comment / sample

Raw positive

–

marker+ / total detected events

Corrected positive

–

after background correction

QC interpretation

–

not a biological diagnosis

Interpretation

Calculating…

Quantitative output

QC risk indicators

Background burden

–

Coincidence risk

–

Detergent control

optional

Thresholds are empirical EV-iTEC guidance values for QC orientation, not validated consensus cut-offs.

Biological caution

Cautious biological interpretation

Models & equations

Marker profile: CD9 / CD63 / CD81 / CD29

Enter background-corrected positive fractions if available. This profile is descriptive and intended to support cautious interpretation, not classification.

CD9

Corrected positive %

CD63

Corrected positive %

CD81

Corrected positive %

CD29

Corrected positive %

Ratios

Ratios use corrected positive fractions. They are robust only when background, gating and antibody performance are comparable.

Profile interpretation thresholds (e.g. CD29 prominence, tetraspanin spread) are empirical EV-iTEC guidance values, not validated consensus cut-offs. They are intended for QC orientation only.

Cautious profile interpretation

Optional replicate summary

Paste one corrected positive fraction per biological replicate. Use percentages or fractions; both are accepted. Example: 21.5, 23.1, 19.8

Corrected positive fractions

Marker / condition label

CI uses t-distribution (df = n−1) for n ≤ 30; z = 1.96 for n > 30. Calculated on untransformed proportions — treat as descriptive only, especially for values near 0% or 100%.

–

biological replicates

Mean ± SD

–

descriptive variability

95% CI

–

t-based, approximate

Replicate interpretation

Important note: The output is QC-oriented guidance. Marker positivity, ratios and Poisson-derived λ values are statistical and operational approximations. They should not be interpreted as absolute EV identity, absolute antigen copy number or direct biological function. Biological interpretation requires appropriate controls, orthogonal validation and experimental context.