Defensive duties were mounted along the Libyan border in early 1941, before the brigade was committed to the Syria-Lebanon campaign, fighting against Vichy French forces.

The brigade's constituent units were spread across various locations in Western Australia including Kalgoorlie, Leonora, Boulder, Coolgardie, Albany, Wagin and Northam.

[7] Following a brief period of rudimentary training in home locations in Australia,[8] under the command of Brigadier Jack Stevens, the brigade embarked for the Middle East in October 1940 aboard the Aquitania.

[18] On 6 July, the 2/16th Battalion attacked the main Vichy defensive position on El Atiqa Ridge,[19] launching a frontal assault on a force of French Foreign Legion troops.

[22] In December 1941 Imperial Japanese forces advanced rapidly in South East Asia following the attack on Pearl Harbor and the invasion of Malaya.

[23] In February 1942, however, as the situation in the Pacific worsened, the Australian government decided that the division should be brought back to Australia rather than be committed to the fighting in Java, or in Burma.

After a period of leave, the brigade was reconstituted and moved to northern New South Wales and then Queensland where they undertook a defensive role around Caloundra in case of a Japanese invasion.

[25] Under the command of Brigadier Arnold Potts, who had taken over from Stevens in April,[26] they were sent to Port Moresby from where they reinforced the militiamen of the 39th Battalion, fighting a rearguard action on the Kokoda Track.

[25] Throughout late August and into September the brigade fought a number of delaying actions along the track, initially around Isurava, but as the Japanese advance towards Port Moresby continued they were pushed back to Ioribaiwa.

[31] In late November, the brigade, under the command of Brigadier Ivan Dougherty who had replaced Potts in controversial circumstances following the withdrawal from Kokoda,[32] was committed to the fighting around Gona.

[37] Following this, the 21st Brigade headquarters moved to the Huggins Road Block area while its three infantry battalions, which had all been reduced to below company-strength due to losses, began the process of "mopping up" around the coast to the west of Gona.

[40] By that time the brigade was down to about 44 per cent of its authorised "jungle" establishment of 119 officers and 2,415 other ranks,[41] and subsequently it underwent a period of rebuilding around Ravenshoe, in Queensland in preparation for further operations.

[47] Supported by 'B' Company, 2/1st Machine Gun Battalion,[48] the 21st Brigade quickly reached their initial objectives and the following day began to advance towards the east, arriving at the airfield at Sepinggang before noon on 2 July.

On the third day of the fighting, the 21st Brigade came up against determined Japanese resistance after crossing the Batakan Ketjil river but this was overcome in the afternoon following a naval barrage.

[49] The next day they resumed their advance to the east but were subsequently engaged and held up by Japanese coastal defence artillery near the Manggar Besar river.