Zooplankton and zooneuston observations were made at seven stations (four from the Gulf of Aqaba and three from the northern Red Sea), during September and October 2016. The main objective of this study was to assess the variability of nycthemeral fauna in relation to the sampling methods using two different types of nets namely, WP2 net and Neuston net along the two study sites, i.e., the Gulf of Aqaba and the northern Red Sea. Zooplankton was sampled vertically using a standard WP2 net from a depth of 200 m to the surface, whereas zooneuston was made using a standard Neuston net from a depth of 0–10 cm of the water surface. Total zooplankton density was maximum during night time ((617.83 ± 201.84) ind./m3) at the Gulf of Aqaba and total zooneuston was maximum during night at the northern Red Sea ((60.94±29.48) ind./m3), respectively. The most abundant taxa were Copepoda, Gastropoda, Bivalva, Chaetognatha, Tunicata and Ostracoda. The abundance was almost 50% higher at night time at both the Gulf of Aqaba and the northern Red Sea. Overall, 30 taxa covering 10 phyla and 27 taxa covering 8 phyla were recorded in the Gulf of Aqaba and the northern Red Sea. Present work is a preliminary attempt to study the variability of nycthemeral and spatial zooplankton and zooneuston diversity and abundance in the Gulf of Aqaba and northern Red Sea regions, which necessity the need for more extended investigations to understand the local and large-scale processes of this complex, but unique ecosystem.