The Alamire scriptorium calligraphed and illuminated a sumptuous chanson album for the mournful regent Marguerite of Austria. Though its pages are studded with "regret" chansons, the manuscript opens with an uplifting Marian motet, a six-voiced Ave Sanctissima Maria. The paraliturgical text, close to that set by Isaac, lauds the Blessed Virgin in seven phrases (a number itself symbolic of her); the music employs strict canon throughout, deriving six voices from three. The composer manipulates his canonic textures masterfully, following the structure of a textbook Ciceronian oration, as did Josquin in his Ave Maria. The opening phrase, hailing "holiest Mary," transpires in scintillating and full six-voiced textures. The distance of canonic response then expands, and the body of the motet presents antiphonal alternation between the three leading voices and the three answering; as the canonic trio is placed a fourth higher, this creates a continual upwards aspiration within the texture. At the text "In this I do not doubt," a more solid and deliberate bass line prepares for the climactic return of all six voices in the closing prayer; the vocal space also broadens to its widest span in the peroration. Motivic entries throughout feature a descending third and oscillation between B flat and B natural; these last moments retain the cross-relations, yet reverse the melodic motive. All is thus glittering ascent.

A later printed source gives this motet a conflicting attribution to Philippe Verdelot, but scholarly consensus awards it to La Rue; he chose it as model for his own Missa Ave sanctissima.

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