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Abstract Background Drug checking services aim to provide compositional information for the illicit drug supply and are being employed in public health responses to extreme rates of overdose associated with fentanyl within street opioids. The technologies used within these services range from basic qualitative tests, such as immunoassay test strips, to comprehensive quantitative analyses, such as mass spectrometry. In general, there is concern that heterogeneity of a drug mixture adds significant uncertainty when using drug checking results based on a small subsamples. The presence of hot spots of active drug components in this context is often termed the ‘chocolate chip cookie effect’. Establishing the limitations of the service are essential for interpretation of the results. Methods This study assesses the consequence of drug heterogeneity and sampling of consumer level opioid purchased in Victoria, British Columbia ( $$n=21$$ n = 21 , 50–100 mg each) on quantitative fentanyl results determined from testing with paper spray mass spectrometry. Results: Using descriptive statistics, such as relative standard deviation and interquartile range, the results demonstrate varied distributions of fentanyl concentrations within a single drug batch. However, the presence of hot spots, defined as outliers, were relatively rare. Conclusions This study found that the variability in fentanyl concentration from drug heterogeneity and sampling is greater than that attributed to the analytical technique. On a practical level, this provides data to help guide communication of limitations of drug checking services, supporting the aim of trust and transparency between services and people who use drugs. However, if drug checking services continue to be restricted from fully engaging with the reality of manufacturing, buying, selling, mixing and dosing practices, the accuracy, usefulness, and impact will always be limited.