This review summarizes our current knowledge and understanding of the biosynthesis and assembly of lipoprotein(a) [Lp(a)] with a focus on diagnostic and therapeutic implications. Lp(a) is composed of a low-density lipoprotein (LDL)-like particle and the covalently bound apolipoprotein(a) [apo(a)]. Our understanding of the physiology and pathophysiology of the atherogenic Lp(a) has considerably increased over the past decades. The precise mechanisms regulating the biosynthesis, secretion, and assembly of Lp(a) have been extensively investigated but are, nevertheless, still incompletely understood or, at least, controversially discussed. Lp(a) plasma concentrations are mainly determined by synthesis and, to a minor extent, also by catabolism. Assembly of Lp(a) occurs in a complex 2-step procedure, starting with an intracellular non-covalent association between lysine-binding sites on apo(a) and lysine residues on apoB-100, the two major protein components of Lp(a). The final assembly of Lp(a) occurs extracellularly and/or transiently associated with the cell membrane by covalent disulfide bonding from newly synthesized apolipoprotein(a) and circulating LDL. Lp(a)-lowering therapies have been developed that specifically inhibit apo(a) biosynthesis and Lp(a) particle assembly. The inhibition of Lp(a) assembly raises interesting questions regarding the quantification of Lp(a) since it leads to the release of substantial amounts of LDL-unbound apo(a), which is co-measured by routine laboratory assays. The complex biosynthesis and assembly pathway of Lp(a) has been intensively investigated since its discovery more than 60 years ago. It is not only of basic-scientific interest but also concerns actual issues of diagnosis and therapy of elevated plasma concentrations of this enigmatic lipoprotein.