No. 121 Squadron was formed 14 May 1941 at Kirton-in-Linsey in 12 Group and assigned the squadron code AV. Peter R. Powell, a veteran of the Battle of Britain credited with destroying 7 german planes, was picked to be the first squadron leader. Royce Wilkinson and Richard Moore transferred from No. 71, Bob Reed, Collier Mize, and Fred Scudday from No. 43, and Loran Laughlin and Carrol McColpin from No. 607. Most of the rest of the pilots were drawn from operational training units or OTUs. During the initial 2 months training No. 121 flew the Hurricane I and when the squadron was declared operational on 21 July 1941 had transitioned to Hurricane IIb's. Encounters with enemy aircraft were minimal, the first contact coming on 8 August 1941 when Sel Edner and Jack Mooney damaged a Ju 88. On 18 August Peter Powell probably destroyed a Me 109F when the squadron was patroling between St. Omer and Dunkirk. For the remainder of 1941 the squadron flew primarily convoy patrol missions with the occasional sweep or escort duty into France from staging areas in 11 Group.
Fred Scudday and Joe Durham saw thier first action when on a routine convoy patrol they spotted a Ju 88 making a run on the convoy. Scudday wrote: "The pilot must have spotted us before we discovered him. He had turned away from his attack on the ships and was flying out to sea. I gave Fred a few seconds after he started his dive to attack, to make certain he would be clear of my guns. Then I made my pass. Scudday's first burst of fire tore away the top turret of the bomber and caused the port engine to explode in a ball of smoke and flame. There was no question that the Junkers was already mortally wounded when I pressed the firing button for my guns and added the coup de grace. As I made my climbing turn to rejoin Fred I noticed that the enemy bomber had vanished into the sea, without a trace." Jim Griffin said of convoy patrol:
"We were flying at an altitude of nine hundred feet, just beneath the clouds, and had been circling the ships for a half hour. It would be another hour before we'd be relieved by four other 121 Eagle Squadron pilots. The weather was always treacherous, and there was never anything between the ships and enemy territory but a stretch of open water. Convincing yourself that flying endlessly between a hostile sky and deadly sea was boring was better than listening to the nagging voice of your subconscious reminding you of the unpleasant features of such a patrol: the fact that survival time in the icy water below would be a matter of minutes; that engine failure beyond gliding distance to land would leave you with two grim choices: ride the Spit down to what would be a quick merciful end... or bail out, knowing that even if there was enough altitude for your chute to open and lower you gently into the sea you would probably perish before help could arrive."In October the squadron received Spitfire IIa's and in November changed to the Spitfire Vb. On 16 December the squadron was transferred to North Weald, in 11 Group, and in mid-January Hugh Kennard became squadron leader.
Bad weather early in 1942 hampered flight operations but No. 121 did take part in the acton against the German battleships Gneisneau and Scharnhorst and the cruiser Prinz Eugene on 12 February. During the second week of March the unit engaged in combat against German aircraft while patrolling over France and on 23 March got it's first kill when a FW 190 was destroyed by Jack Mooney near Calais. Reade Tilley was credited with probably destroying another FW 190. In April things started to heat up with the squadron flying 740 operational missions and claiming 4 destroyed, 1 probable, and 3 damaged, at the cost of 3 pilots. On 12 April Leroy Skinner destroyed a FW 190 with Kennard, Barry Mahon and Thomas Allen getting credit for damaging german planes. On 15 April Skinner and Sel Edner each destroyed an FW 190 and Allen damaged one. On 24 April Skinner shared credit with Jim Daley in the downing of a Ju 52. On 17 May Jim Daley and Sel Edner each downed an enemy plane. In late May the focus of the squadron shifted to anti-shipping work with the group combining to sink an armed trawler on 20 May, Jim Daley sinking a 1,000 ton Minsweeper, as well as downing an enemy plane on 27 May, and another flight jointly sinking a 2,000 ton Minesweeper.
June was a busy month for No. 121 with the squadron flying sweeps, escort, and Rhubarbs, and moving to Southend-on-Sea on 3 June. On 9 June 4 FW 190's were destroyed over St. Omer, Barry Mahon and John Mooney getting 2 each. No. 121's best day came on 31 July with 7 destroyed over Abbeville, France. In this engagement Selden Edner and Barry Mahon each destroyed 2 and Squadron Leader Hugh Kennard, Sgt/Plt William Kelly, and Frank Boyles destroyed one each. Kennard was wounded so another Englishman, W. Williams took over command. No. 121 flew three sorties during the Dieppe raid of 19 August and claimed 1 destroyed, 2 probable and 1 damaged FW 190's. This was the last intense action 121 would see as Eagles and by the time they disbanded they were credited with 18 enemy planes destroyed. They were transferred to the USAAF on 29 September, where they became the 335th Fighter Squadron of the Fourth Fighter Group.