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Introduction: Globally, COVID-19 pandemic has a significant impact on mental health. In Nepal, COVID-19 positive cases have to self-isolate at home in multi-generational and multi-family households. This could be strongly associated with depression, anxiety, and stress-related health outcomes. Additionally, COVID-19 related stigma and fear of transmission may intensify depression, anxiety, and stress symptoms. This study determined the prevalence of depression, anxiety, and stress symptoms and their association with presence of COVID-19 symptoms and comorbid conditions among home isolated COVID-19 positives in the Karnali province, Nepal. Methods: We conducted a cross-sectional study to assess depression, anxiety, and stress symptoms among 402 home isolated COVID-19 patients of Karnali province from January to May 2021 using “Depression, Anxiety and Stress Scale-21 (DASS-21)”. We interviewed patients to collect socio-demographic, DASS-21, COVID-19 symptoms, comorbid conditions, and self-treatment. We conducted a telephonic interview using a standardized questionnaire using Kobotoolbox. We calculated the prevalence of depression, anxiety, and stress symptoms. We utilized univariate and multivariate logistic regression to determine their association with the presence of COVID-19 symptoms and comorbid conditions. In multivariate logistic regression, we adjusted sociodemographic factors (age, gender, ethnicity, marital status, monthly family income, education level), smoking status and history of self-treatment. We reported adjusted odds ratios (aOR) with 95% confidence intervals. All analyses were conducted in R (version: 4.0.3). Results: The prevalence of depression, anxiety and stress symptoms among home isolated COVID-19 patients were 8.0% (95% CI: 5.5 to 11.1), 11.2% (95% CI: 8.3 to 14.7), and 4.0% (95% CI: 2.3 to 6.4) respectively. Higher odds of depression symptoms (aOR: 2.86; 95% CI: 1.10–7.44, p = 0.03), anxiety symptoms (aOR: 3.81; 95% CI: 1.62 to 8.93; p =