Quantification of the α-subunit of ammonia monooxygenase (amoA) through PCR is an established technique for estimating the abundance of ammonia oxidizing archaea (AOA) in environmental samples. This study quantified AOA with two established primer sets in 1 cm increments from the sediment surface (0-1 cm) to a depth of 10 cm at two locations within Port Phillip Bay (PPB), Australia. Primer choice had a significant effect on within sample estimates of AOA with copy numbers ranging from 102 to 104 copies per ng DNA. Variation in AOA abundance patterns with increasing sediment depth were site and primer specific. Sequence mismatches between the primer binding region of the isolated amoA sequences from PPB and Nitrosopumilus maritimus SCM1 were identified and may explain the high variation identified between primer estimates. Our results highlight the need for testing multiple primer pairs that target different regions of the AOA amoA sequence prior to large-scale marine sediment environmental studies.