In birds, the number of sperm trapped between the perivitelline membranes around the ovum is an estimate of sperm numbers present at the time and place of fertilisation in the female reproductive tract. Sperm numbers may vary among species and between eggs in a clutch and can provide information about sperm utilisation and mechanisms of sperm competition. Here, we describe patterns of variation in sperm numbers through the egg-laying sequence in three passerines in which extra-pair paternity is common, but copulation behaviour differs. Sperm numbers showed no systematic change across the laying sequence in blue tits (Cyanistes caeruleus), but decreased significantly with laying order in bluethroat (Luscinia svecica) and tree swallow (Tachycineta bicolor) clutches. This is consistent with observations that blue tits regularly copulate throughout the laying sequence, while bluethroats stop mate guarding and tree swallows reduce their copulation frequency once the first egg is laid. Nevertheless, cases of a sudden increase in sperm numbers in clutches of bluethroats and tree swallows suggest that successful inseminations also occurred after laying started. In blue tits and bluethroats, sperm numbers were not higher on extra-pair sired eggs than on eggs sired by the social male, suggesting that extra-pair copulations are not timed to the period of peak fertility for each egg. More extra-pair offspring originated from eggs laid early in the sequence in blue tits, while there was no systematic bias in bluethroats. Our results suggest that copulations during the laying sequence are predominantly performed by within-pair males in our study species.