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Measurement of specific loss power (SLP) of magnetic nanoparticles is crucial to assert the heating potential in magnetic hyperthermia. There has been a significant improvement in characterizing magnetic nanoparticles’ heat-triggered functions by many research groups. However, optimal experimental conditions along with notable determination methods of the SLP in magnetic hyperthermia have not been widely proposed until now. Despite the remarkable progress in this field, the evaluation process of SLP suffers from uncertainties and errors imposed not only by experimental parameters (depending on the particles, the conditions and the measurement) but by the estimation methodology, as well. In this work, we propose a step by step standardization protocol, starting from definition and recording of potential uncertainty and error sources, present during a typical magnetic hyperthermia experimental protocol. The error of each specific parameter is estimated and translated to ultimate heating efficiency evaluation. According to our analysis, magnetic hyperthermia experiment and its corresponding estimation, under non-adiabatic conditions, may lead to a propagated uncertainty up to 14% on the SLP value. Meanwhile, different heating evaluation methods were assessed under a wide range of experimental conditions, with the ‘modified law of cooling’ proving to be the most accurate one—limiting the SLP uncertainty to values under 5%—compared to the ‘initial slope’ and ‘Box–Lucas’ methods, which show a remarkable uncertainty of over 15%. All parameters involved in the heating efficiency evaluation and their associated uncertainties analysis presented in this work, are included in a standardisation protocol, a handy guideline for determining accurate, reliable and reproducible SLP values, thus adequately evaluating its impact in potential bioapplications. [ABSTRACT FROM AUTHOR]