Abstract: An experiment was conducted to study the response of three chicken lines with different HSP 70 genotypes to acute heat stress. Twenty eight kampong chicken (native chicken, with seven genotypes i.e., AA, AB, AC, CC, AD, DD and BC) and twenty four Arabic chicken (with six genotypes i.e., AA, AB, AC, CC, AD and BC) and four commercial chickens (with one genotype i.e., DD) were used and randomly allocated in a factorial arrangement into groups which their HSP 70 genotypes had been identified. Acute heat stress was exposed at 40°C for 0.5, 1.0 and 1.5 h, respectively, using chamber in 33 x 33 x 75 cm3. Parameters measured were the onset of panting (minute), panting frequency (times/minute), feed consumption (g/minute), water consumption (mL/minute), manure water content (%), rectal temperature (°C), serum corticosterone concentration (μg/dL) and HSP 70 expression (copy mRNA). The result of this study showed that there was an interaction between chicken lines and acute heat stress exposure on water consumption and manure water content. Chicken lines affected panting frequencies and manure water content but it did not affect the onset of panting, feed and water consumptions, rectal temperature, serum corticosterone concentration and expression of HSP 70. Acute heat stress increased panting frequency, drinking water consumption, manure water content, rectal temperature, serum corticosterone concentration, HSP 70 expression and it decreased feed consumption. The highest response on panting frequency, rectal temperature, serum corticosterone concentration and expression of HSP 70 was found in the DD genotype and the lowest in AD genotype. The most rapid onset of panting occurred in DD genotype and the slowest in AD genotype. The study revealed interaction between chicken lines and HSP 70 genotypes in heat resistance. Kampong chicken had the highest heat resistance as compared to Arabic and commercial chickens but HSP 70 genotypes that was the most tolerant to high ambient temperature was AD genotype where as the lowest tolerant was DD genotype.