Natural autoantibodies (NAAbs) are immunoglobulins generated in the absence of antigen provocation, with no inherent pathogenic effect, but instead involved in maintaining immune homeostasis. We aimed to evaluate the implication of immunoglobulin G (IgG) antibodies against heat shock protein 90α (HSP90α) in systemic lupus erythematosus (SLE) patients, focusing on how those NAAbs may interfere with the immunothrombosis process. Anti-HSP90α IgG was measured by an enzyme-linked immunosorbent assay in the plasma of 308 SLE individuals, paired with 308 healthy controls (HC). Validation was performed in another cohort, comprised of serum from 125 SLE and 77 HC. In an in vitro experiment, platelets were cultured with mitochondria (abundant sources of HSP90α) treated with anti-HSP90α IgG, and the activation markers thrombospondin-1 and platelet factor 4 were measured in the supernatant. Levels of anti-HSP90α IgG were comparable in SLE and HC, but decreased in the SLE patients with secondary antiphospholipid syndrome (APS), both in the exploratory and the validation cohort (p = 0.0265, and p = 0.0472 respectively). Mitochondria triggered platelet activation in a TLR2- and HSP90α-dependent manner (p = 0.0152 and p <
0.002 respectively). Anti-HSP90α IgG emerges as a prominent NAAb-modulating immunothrombosis in SLE and holds promise as a biomarker of APS risk in these patients.