The aquarium is focused on local aquatic animals from the San Francisco Bay and neighboring rivers and watersheds as far as the Sierra Mountains.

[4] Specific objections included the amount of fill required (an additional 24,000 square feet (2,200 m2) would need to be reclaimed from the Bay) and a potential violation of the city's 1990 Proposition H, which prohibits nonmaritime use of waterfront property.

[10] The signature attraction, two long acrylic underwater tunnels with approximately 770,000 total gallons of San Francisco Bay salt water.

The aquarium opened on 19 April 1996[1] under the name UnderWater World at a cost of US$38,000,000 (equivalent to $75,980,000 in 2023),[4] filled with approximately 4,000 fish with 100 unique species indigenous to San Francisco Bay.

[11] After being shown a short introductory film, visitors walk through an area with three pools, then take an elevator down to the signature attraction, which is two acrylic tunnels 360-foot (110 m) long overall that cuts through two tanks filled with total of 707,000 US gallons (2,680,000 L; 589,000 imp gal) of filtered water from the bay, based on a similar transparent tunnel in an aquarium of the same name in Auckland, New Zealand.

UnderWater World responded by cutting ticket prices to US$12.95 (equivalent to $24.58 in 2023), the city planning commission removed the requirement for subsidised parking and buses, and Pier 39 vowed to paint the exterior with murals, create joint marketing opportunities with other San Francisco institutions, and bring in more impressive animals.

[9] UnderWater World was sold to a group led by BNP Paribas in June 2000, who announced plans to rename it to Aquarium of the Bay and add more sea animals in 2001.

[15] The entity's accreditation from the Association of Zoos and Aquariums was revoked, following findings of deteriorated equipment and poor regulation of animal tank water temperature.

The aquarium received funding from the San Francisco Public Utilities Commission (SFPUC) to create an internship program for select high schools.

Within some of the education and outreach programs for schools, the Aquarium of the Bay offers field trips that support Next Generation Science Standards (NGSS).

For 1st through 3rd graders, the aquarium offers a program to teach them about the life cycle of a salmon from egg to adulthood in the San Francisco Bay, Pacific Ocean, and rivers of California.

[17] A local businessman, Darius Anderson, owner of Kenwood Investments, put up the funds to create a competitive bid,[8] with a condition allowing The Bay Institute to purchase it from Kenwood Investments in a few years' time at a predetermined price, which would eventually transform the aquarium's mission from entertainment to a nonprofit education and research center.

[19] Aquarium of the Bay has collaborated with a researcher named Matt Savoca at UC Davis to study olfactory reception in anchovies and plastic uptake.

Along the Embarcadero and other spots around San Francisco, hand-painted, life-size sea lion statues made of carbon fiber can be found.

After the Loma Prieta earthquake hit San Francisco, the lions bravely and mysteriously started to bask in the sun on the K-dock on their pier.

Aquarium of the Bay commissioned 30 local artists to paint unique carbon fiber sea lions in Sausalito before taking them out and placing them in various locations around San Francisco for people to find as an homage to the now long-time residents.