<p>Internalized transphobia (IT) is the rejection that transgender and gender non-conforming people (TGNC) feel towards themselves because of the internalization of negative values, prejudices, and/or stereotypes that prevail in society. This study aimed to adapt and validate a Spanish version of the Transgender Identity Survey (TIS) to assess IT in Spanish-speaking TGNC individuals. The sample consisted of 207 TGNC Spanish adults aged between 18 and 63 years (25.11 ± 8.16) who completed the TIS, along with measures of self-esteem and body image. Translation, cross-cultural adaptation, and validation analyses of the TIS were carried out. A reduced version of the original scale composed of 15 items in a one-dimension to assess IT was obtained through Exploratory Factor Analysis (EFA). Validity evidence based on the internal structure was tested through Confirmatory Factor Analysis (CFA). The indices from the CFA showed that the reduced version of the TIS had an acceptable fit: χ² = 250.11, p &lt; 0.001; CFI = 0.86; GFI = 0.96; RMSEA = 0.09 (0.07–0.10); SRMR = 0.06. The scale showed adequate reliability values (α = 0.890; Ω = 0.892). Correlations between TIS, self-esteem and body image reported convergent validity evidence. The Spanish version of the TIS provides a useful, reliable, and valid instrument for both research and clinical areas, supporting interventions aimed at reducing internalized stigma and promoting well-being and self-acceptance in the gender-diverse community.</p>