We compared the construct stability of traditional knotted techniques with modern knotless anchor systems used in arthroscopically assisted latissimus dorsi transfers (aLDTs) for irreparable posterosuperior rotator cuff tears. Eighteen cadaveric shoulders were age- and gender-matched to the following three groups: knotted group (two knotted anchors); knotless group (two knotless anchors); and triple anchor group (two knotless anchors; one all-suture anchor at the apex of the humeral head). All tendons were cyclically loaded in line of the aLDT over 400 cycles followed by a load to failure test. Outcome measures were the ultimate load to failure, elongation, construct stiffness, and failure modes. The triple anchor group revealed the highest ultimate load to failure (431 ± 78 N) compared with the knotted (326 ± 52 N; p = .022) and knotless (353 ± 105 N; p = .129) groups. Total elongation and construct stiffness were not significantly different comparing all three groups. The failure modes were: anchor pull-out in all specimen of the knotted group; three (50%) anchor pull-out and three suture pull-out in the knotless group (p = .046); four (67%) anchor pull-out, one (17%) suture pull-out and one tendon pull-out in the triple anchor group. Biomechanical analyses of knotless fixation techniques for aLDTs show similar construct stability and elongation compared with the traditional, knotted techniques. Bone fixation seems to be improved with the knotless anchor systems. In our practice, we continue to use the knotless fixation technique for aLDT and in the situation of weak, osteoporotic bone, we add a third (all-suture) anchor to improve construct stability.