For efficient analysis and characterization of biological ice nuclei under immersion freezing conditions, we developed a Twin-plate Ice Nucleation Assay (TINA) for high-throughput droplet freezing experiments, in which the temperature gradient and freezing of each droplet is tracked by an infrared detector. In the fully automated setup, a couple of independently cooled aluminum blocks carrying two 96-well plates and two 384-well plates, respectively, are available to study ice nucleation and freezing events simultaneously in hundreds of microliter range droplets (0.1–40 µL). A cooling system with two refrigerant circulation loops is used for high-precision temperature control (deviations < 0.5 K), enabling measurements over a wide range of temperatures (~ 233–270 K) at variable cooling rates (up to 10 K min −1 and more). The TINA instrument was tested and characterized in experiments with bacterial and fungal ice nuclei (IN) from Pseudomonas syringae (Snomax ® ) and Mortierella alpina , exhibiting freezing curves in good agreement with literature data. Moreover, TINA was applied to investigate the influence of chemical processing on the activity of biological IN, in particular the effects of oxidation and nitration reactions. Upon exposure of Snomax ® samples to O 3 and NO 2 , the concentration of IN active around 269 K (−4 °C, “class A”) decreased by about two orders of magnitude, while the concentration of IN active around 265 K (−8 °C, “class C”) decreased by about one order of magnitude. Furthermore, TINA was used to study aqueous extracts of atmospheric aerosols, simultaneously investigating a multitude of samples that were treated in different ways to distinguish different kinds of IN. For example, heat treatment and filtration experiments indicated that highly efficient biological IN were smaller than 0.1 µm, and many IN active between 263 K (−10 °C) and 256 K (−14 °C) were heat-resistant and larger than 5 µm. The results confirm that TINA is suitable for high-throughput experiments and efficient analysis of biological IN in laboratory and field samples.