One of the main factors that affect animals’ performance is high temperature, causing several changes, including at the cellular level. These changes lead to an increased expression of heat shock proteins. This experiment aimed to study the HSPA1A and HSP90AA1 gene expressions of Holstein cows after exposure to direct solar radiation. The heat tolerance test was performed and rectal temperature and respiratory rate measured, and blood samples collected. After the erythrocytes lysis to obtain the buffy-coat, the RNA was isolated by the TRIzol method and RT-PCR performed with SuperScript III after digestion with DNase I. The qPCR apparatus took place in 7500 Real Fast Time, using TaqMan Gene Expression Assays for HSPA1A and HSP90AA1 target genes, ACTB and PPIA as endogenous genes. The ΔCt (Ct_target − Ct_endogenous) were calculated as well as gene expression through the 2^-ΔΔCt method. The treatments considered for statistical analysis were t < 44°C and T ≥ 44°C (with T being the black globe temperature). We used linear mixed models in the program R Project Software (version 3.0.1). There weren’t significant differences between treatments for any of the variables. This way we can say that animals were in a moderate stress condition, which did not allow the identification of differences in HSPA1A and HSP90AA1 gene expression.