The Los Angeles gallery has imposed the strict time limit on Infinity Mirrors, a series of six installations by the Japanese artist, in a bid to keep people funnelling through quickly.
Vistors will have to ensure they take any photographs within their 30-second window, before being moved on.
The six Infinity Mirror Rooms are small boxes that use light and mirrors to create visual trickery.
The so-called "selfie rule" is expected to prevent these spaces becoming too crowded – avoiding accidents like the one last year, which saw one of Kusama's iconic pumpkins smashed by a clumsy visitor trying to take a picture of themselves.
It should also increase the amount of people able to visit the exhibition without one of the 90,000 advanced tickets, which sold out in a matter of hours.
But according to Artnet, those invited to the exhibition's preview last week were "stressed" by the new rule.
"I got a little stressed because you had like one second to take pictures," said a visitor when stepping out of The Souls of Millions of Light Years Away artwork.
During its time at London's Victoria Miro gallery last year, Kusama's Infinity Mirrors exhibition was endlessly shared on Instagram by visitors.
The gallery has now created a specific Instagram location tag for Kusama's exhibition, which is already filled with pages of selfies.
Although Kusama's work has gained a much larger audience in recent years, the artist has been practising since the 1960s.
She famously creates all of her work in a studio near the Tokyo psychiatric facility in which she has lived, voluntarily, since 1977, having reported experiencing visual and auditory hallucinations her whole life.
Prior to her admission, she spent a period of time living and working in New York City, where she was part of the avant-garde art scene.
Her life and works have recently been immortalised in a children's book by illustrator Ellen Weinstein and MoMA curator Sarah Suzuki, which documents her journey to success through painterly illustrations.
Yayoi Kusama: Infinity Mirrors opened to the public on 21 October 2017, and is on show at The Broad until 1 January 2018.