Binocular indirect ophthalmoscopy (BIO) is the gold standard for retinopathy of prematurity (ROP) screening, however, associated use of an eyelid speculum results in significant infant stress. Handheld optical coherence tomography (OCT) may identify ROP severity biomarkers and is non-contact. We performed a prospective, observational study to compare vital signs measured during BIO and OCT to identify whether OCT has an advantage in lowering cardiorespiratory impact of screening. This study included 16 premature infants screened for ROP, recruited between April 2019 and February 2020. Each infant underwent BIO and OCT at least 30 minutes apart, alternating the order at each consecutive screening session. Infants who were too unstable were excluded from the study. Vital signs were obtained 1 minute before, 1 and 2 minutes into, and 15 minutes after the examination. Vital sign deviations from baseline were compared for each infant between the two imaging modalities using a paired linear mixed model to adjust for multiple imaging sessions.This study included 26 examinations among 16 infants (62.5% female, mean gestational age 285.86±2.82 weeks, mean birth weight 1058.25±289.00 grams) with 9 infants at ROP stage 1, 4 at ROP stage 2, and 1 at ROP stage 3. BIO duration was significantly shorter than OCT (4.4 vs.10.8 minutes, P<0.001). Vital signs increase from baseline were significantly greater for BIO compared to OCT for heart rate at 1 minute (18.81±20.75 vs. 0.00±22.82 beats per minute, P=0.04), diastolic blood pressure at 2 minutes (41.22±31.69 vs. 28.19±25.84 mmHg, P=0.04), mean arterial pressure at 2 minutes (46.83±34.83 vs. 36.99±20.08 mmHg, P=0.04), and systolic blood pressure at 15 minutes (10.85±10.55 vs. -5.12±13.53 mmHg, P=0.04). OCT resulted in significantly lower impact on heart rate and blood pressure compared to BIO. The ability to screen for ROP using OCT may benefit overall health for these vulnerable premature infants.