Yellow leaf disease (YLD), caused by sugarcane yellow leaf virus (SCYLV), has been reported to infect sugarcane worldwide causing significant yield losses and is considered a major disease of this crop. A panel composed of 98 genotypes encompassing basic germplasm, commercial cultivars, and elite clones was assayed in the nursery and a replicated field for resistance to SCYLV, combining symptom expression and virus quantification. Virus symptom intensity was evaluated using a diagrammatic scale while virus titer was estimated by DAS-ELISA (Double Antibody Sandwich-ELISA) and RT-qPCR (Reverse Transcription Quantitative PCR) in the nursery and field trials, respectively. Resistance was evaluated alongside symptom development kinetics. Based on the symptom intensity, 52 (53.06%) genotypes were classified as resistant in the nursery and 42 (42.86%) in the field. Twenty-nine (29.59%) genotypes showed no symptoms in the nursery and field trial. Moderately resistant genotypes showed a low correlation between virus titer and symptom intensity. The kinetics of symptom development increased over time in moderately susceptible and susceptible genotypes. The SCYLV incidence assessed by RT-qPCR was 92.55%, which was detected in 83% of the asymptomatic genotypes. The broad-sense heritability based on symptom expression and relative quantification was 52.62% and 68%, respectively. The virus quantification assays allowed for the identification of potential genotypes immune to SCYLV. •Resistance evaluation combining symptom expression and virus quantification.•83% of asymptomatic but SCYLV infected genotypes.•Five potential SCYLV immune genotypes identified by DAS-ELISA and RT-qPCR.•Combination of RT-qPCR and symptom expression proven suitable for parental selection.•Moderate to high broad-sense heritability for symptom expression (52.62%) and virus titer (68%).