<p>This work focuses on the coupling effect of cooling rate (CR) and inclusion characteristics on intragranular acicular ferrites (IAFs) nucleation and growth for Al–Ti–Ca deoxidized high-strength low-alloy steel through <i>in-situ</i> observation with high-temperature confocal laser scanning microscope (HT-CLSM). The results indicate with increasing CR from 0.5 to 7 °C/s, the initial transformation temperature of ferrite and the finishing temperatures of microstructural transformation lower, and the difference of initial transformation temperatures between IAFs and ferrite side plates (FSPs) decreases. As CR is increased to 7 °C/s, FSPs, IAFs and intragranular bainitic ferrites (IBFs) nucleate almost simultaneously at 560.1 °C to 551.0 °C. For inclusion characteristics, increasing CR from 0.5 to 7 °C/s doesn’t alter inclusion types, but reduces their size from 1.74 to 1.25 <i>μ</i>m and increases their number density from 143.42 to 377.22 mm⁻². Notably, the complex inclusions of CaO–Al₂O₃–Ti₂O₃+(Ca,Mn)S, as the most effective type for inducing IAFs formation, increase in their number density, and decrease in their average size with increasing CR. But on their surface, the uneven distribution of Ti-oxides increases and (Ca,Mn)S content reduces, resulting in a decrease in their nucleation ability for IAFs. In addition, the actual growth rate of IAFs increases from 14.62 to 44.88 <i>μ</i>m/s with the increase of CR, but the area fraction of IAFs initially increases and then reduces, reaching a maximum value of 67.3&#xa0;pct at 1 °C/s. Meanwhile, HAGB proportion and average GND density also first increase and then decrease with increasing CR, having the maximum value of 76.1 pct and 3.51&#xa0;×&#xa0;10¹⁴&#xa0;m⁻² at 1 °C/s. Therefore, the CR of approximately 1&#xa0;°C/s is recommended to obtain an optimal IAFs microstructure.</p>