Wax-coated dried-milk-spot paper for human milk protein compositional analysis

Studies exploring human milk are often hindered by laborious and costly conventional sampling, resulting in small sample sizes. Here, we implement a paper-based sampling method, dried milk spots (DMS), as an alternative to increase sampling capacity and frequency, primarily tailored for protein compositional analysis. Uncoated paper was unsuited for DMS, with β-casein recovery at 64 ± 1 % and α-lactalbumin at 85 ± 2 % after storage of 14 days. A bespoke wax-coated paper was thus developed to improve protein recovery, alongside an ethanol pre-wetting step to enhance wicking of milk. Using this ethanol-pre-wetted-wax-coated paper, β-casein recovery increased to 80 ± 6 % and α-lactalbumin to 82 ± 1 %, with the β-casein-to-α-lactalbumin recovery ratio reaching 97 ± 8 % after storage of 14 days. This DMS method requires only room temperature storage, facilitates easy transportation, and supports protein compositional analysis, thus holding promise for future application in human milk studies.