my $matched;
while (my $line = <FILE>) {
    my ($one,$two,$three,$four,$five) = split "\t",$line;
    if ($three eq $checkthree) {
        print "<p>With matching parameters I should be able to see this </p>";
        $matched = 1;
        last;
    }
}

unless ($matched) { # this is like the else
    print "<p>If I put the else statement in it ignores the fact that the parameters match - and comes here!  (But I don't know why).</p>";
}