Hypoglycaemic disorders may be classified according to their pathogenesis, which can be insulin mediated or non-insulin mediated. In apparently healthy individuals, hypoglycaemia is more likely due to endogenous hyperinsulinism which could be caused most frequently by an insulinoma and typically occurs in the fasting period, but could also derive from non-insulinoma pancreatogenous hypoglycaemia syndrome (NIPHS), insulin autoimmune hypoglycaemia, post bariatric surgery (dumping syndrome), accidental or fictitious hypoglycaemia (intake of beta cell secretagogues such as sulfonylurea). Idiopathic reactive hypoglycaemia (IRH), which occurs in the post-prandial state, has been associated with delayed and excessive second phase insulin response, an increased insulin sensitivity, or imbalance in incretin secretion, although underlying mechanisms are still not completely clear [
10,
20,
21]. Conditions causing non-insulin mediated hypoglycaemia include states of critical illness, cortisol or GH deficiency, drugs, and certain malignant disorders (through production of IGF-I, IGF-II or pro-IGF-II) [
10,
22]. In patients presenting with a history of hypoglycaemia and documented Whipple triad, diagnosis can usually be established by a laboratory assessment at the time of the spontaneous hypoglycaemic episode. When it is not observed by clinicians, a prolonged supervised fasting test, which can last up to 72 h, has been established as the gold-standard method for the assessment of the role of insulin secretion pattern in hypoglycaemia occurring in the fasting period. Through several series of cases of insulinoma, the Mayo clinic experience [
14] concluded that the 72 h test was necessary to exclude false negative results in the few patients diagnosed on the 3
rd day of the fast. Nevertheless, a shorter modality of the test has been proposed by different authors, due to its practicality and potential first-line screening test: Hirshberg and colleagues [
16] demonstrated that close to 100% of patients with endogenous hyperinsulinemia were diagnosed in less than 48 h. In their study the fast was terminated when plasma glucose was < 45 mg/dL in 43% patients within 12 h, in 67% within 24 h and in 95% within 48 h. Frajans e Vinik (1989) [
17] evaluated 82 patients with endogenous hyperinsulinism caused by beta cell disorders: in blood samples obtained following a 12-h overnight fast in several different days, 76% patients showed glycaemia ≤ 50 mg/dL and in 86% ≤60 mg/dl. Vezzosi and colleagues [
18] applied the same protocol: in their study, 88% of the subjects showed glycaemia < 60 mg/dL, concluding that it could be used as triage do exclude subjects without endogenous hyperinsulinemia. Moreover, Felicio and colleagues [
19] achieved 100% of sensitivity in the diagnosis of hyperinsulinemic hypoglycaemia in blood samples of an overnight fast in 3 consecutive days. In our series, instead, 83.3% of insulinoma patients were diagnosed within 14 h and 91.7% within 24 h. In comparison to the above-mentioned protocols alternative to the gold standard 72 h fast, our protocol has the advantage of being potentially safer since it does not require leaving home after an overnight fast in several days to perform laboratory testing, instead it proceeds the overnight fast on one occasion in a monitored setting (Day Hospital) where hypoglycaemia can be promptly identified and reversed. Our results showed that the overnight fasting glycaemia was significantly lower and corresponding serum C-peptide significantly higher in the insulinoma group, (
p < 0.001 and
p = 0.023) as expected, which is in accordance with the current literature [
1,
4]. The difference in baseline serum insulin, although being higher in the insulinoma group did not reach statistical significance (
p = 0.055). Our preliminary results indicate that the short fasting test when portrayed up to 24 h could reach an excellent sensitivity for diagnosis of insulinoma (91.7%). False positive results deriving from other causes of endogenous hyperinsulinemia as insulin antibodies or NIPHS were not observed. It is also important to consider that during the short fasting test the biochemical evaluation starts at baseline after the overnight fast, 12 h from the last meal, therefore it is not possible to access glycaemia and insulinemia in the preceding time period. Given the likelihood that some patients show diagnostic values earlier, the real time to diagnosis is expected to be even shorter. Furthermore, there was a higher prevalence of the female sex in the insulinoma group (91.7%) in comparison to the non-insulinoma group (76.0%) without reaching statistical significance (
p = 0.38). In current literature, some studies also reported a slightly higher prevalence of the female sex [
2,
4]. Age and BMI were similar between insulinoma and non-insulinoma groups, which was also in accordance with literature [
1]. Importantly, in consideration of the potential risk of dangerous levels of hypoglycaemia in insulinoma patients until arrival to hospital, we observed that the overnight fast was generally well tolerated, and there was no record of complications, which supports the safety of the procedure. In any case, we strongly recommended that the patient monitors symptoms closely and that should be always accompanied by a care giver until admission to the hospital. We do not recommend proceeding with the fast in case of neuroglycopenic symptoms at awakening. Finally, it is necessary to consider the cost-efficacy of both types of tests. The prolonged fasting test requires the availability of a bed for an elective hospitalization for up to 72 h, which in the actual post-pandemic setting, particularly in public hospitals across Europe, is not easily obtainable. The short fasting test reveals to be a more cost-effective alternative, as it is performed in Day Hospital/outpatient setting. The main limitations of this study are the retrospective design and the small sample size.