Purpose: This is the first study to assess the impact of an online microsurgical testicular sperm extraction (mTESE) masterclass. We: 1) describe the masterclass’s scientific content; 2) appraise the participants’ acquisition of knowledge; 3) gauge whether the extent of improvement of the participants’ knowledge/skills was influenced by demographic/professional attributes; and 4) evaluate the participants’ satisfaction. Materials and Methods: This masterclass comprised five didactic lectures followed by 4 case discussions. Online surveys assessed the above objectives using a baseline questionnaire including demographics and past mTESE experience/training, a 24-question pre- and post-quiz, and a satisfaction questionnaire. Results: Participants were between 20–70 years old, with 80.37% males, mainly from Asia, Africa, and Europe, from clinical backgrounds (69.3%), and in public practice (64.4%). Half the sample reported no past mTESE training and very low skills, ≈60% wanted considerably more training, and 50% felt that good training was not readily available. Satisfaction was 98% to >99%. Pre- and post-quiz comparisons confirmed remarkable improvements in knowledge/skills, exhibiting five striking characteristics. Improvements were a) Broad i.e., across 19 of the 24 mTESE questions; b) Deep, of magnitude, as pre-/post-quiz scores improved from mean 13.71±4.13 to 17.06±4.73; c) Highly significant, consistently with p-values <0.001; d) Inclusive i.e., all participants enhanced their mTESE knowledge/skills regardless of demographic/professional attributes; and, e) Differential, e.g., non-clinical/clinical participants improved, but the former improved relatively significantly more, those with ≤5-year experience improved significantly more than those with >5-year, those in public practice significantly more than private practice participants, and those with lower self-rating in performing mTESE significantly more than those with higher self-rating. Conclusions The masterclass was successful with very high satisfaction levels, and markedly improved mTESE knowledge/skills among the participants. Global Andrology Forum’s model can be adopted by organizations with similar goals. Future research needs to evaluate such training to develop a practically non-existent evidence base.