Introduction: The aim of this study was to analyze the choroidal vascularity index (CVI) versus choroidal thickness (CT) as biomarkers in acute central serous chorioretinopathy (CSCR). Methods: In this multicenter retrospective, cross-sectional, noninterventional study carried out at Puerta de Hierro-Majadahonda University Hospital, Madrid, Spain; Jules-Gonin, Lausanne, Switzerland; and Pittsburg University, USA. 40 eyes from 40 patients with acute CSCR, 40 eyes from 40 patients with keratoconus (KC), and 40 eyes from 40 healthy patients were included. The variables analyzed were age, CT, CVI, and the presence of neurosensory retinal detachment. CT and the CVI were obtained from a 12-mm horizontal single-line B-scan (Triton SS-OCT, Topcon Co., Japan). Blinded measurements of the subfoveal CT were performed manually by two independent investigators. The images of the choroid were automatically binarized using a validated algorithm, and a percentage of vascularity was calculated. Results: There were no significant differences in age between the three groups (ANOVA, p = 0.092). There were statistically significant differences in CT and the CVI (ANOVA, p < 0.001). After Bonferroni correction, pairwise analysis between CSCR group against the KC group showed no significant differences in age and CT (p = 0.10 and p = 0.27, respectively). CVI was statistically greater among CSCR patients (p = 0.03). Conclusion: CT does not meet the criteria to be considered a biomarker of acute CSCR, while CVI may prove to be a more specific and reliable biomarker. Further studies with larger sample sizes, standardized procedures, and a wider representation of all CSCR stages are necessary to confirm the validity of CVI as biomarker in this disease. Further studies with larger samples are required in order to validate the use of CVI/CT correlation as a new biomarker.