The president of Jordan's FA, Prince Ali bin al-Hussein, demanded an investigation into whether or not Iranian goalkeeper Zohreh Koudaei is a man.
Koudaei was the hero as she saved two penalties for Iran as they beat Jordan 4-2 in a shoot-out to qualify for the 2022 Women's Asian Cup after their game on September 25 finished 0-0.
It is the first time in the country's history they have reached the finals, with the Iranian manager firmly denying the allegations and insisting Jordan are just looking for excuses.
A spokesman of the Asian Football Confederation (AFC) said: "The AFC does not comment on ongoing investigations and/or proceedings, whether actual or potential."
Koudaei has been forced to defend herself after accusations about her gender previously in Iranian newspapers.
A letter sent by the Jordan FA to Dato' Windsor John, the AFC secretary general, raised doubts about the 'eligibility of a participating player' from the qualifier, played in Tashkent, Uzbekistan.
It alleged the Iranian women's team 'has a history with gender and doping issues' and called for 'due process' to be followed.
Publishing the letter online on Saturday, Prince Ali, a former FIFA vice-president, urged the AFC to 'wake up' to the allegations.
Maryam Irandoost, Iran's head coach, dismissed the allegations against the 32-year-old shot-stopper, who according to the AFC's website previously represented her country in Asian Cup qualifiers in 2008 and 2010.
"The medical staff have carefully examined each player on the national team in terms of hormones to avoid any problems in this regard, and so I tell all fans not to worry," Irandoost told the sports news site Varzesh 3 on Sunday.
"These allegations are just an excuse not to accept the defeat against the Iranian women's national team," she added.
"The Jordanian team considered themselves the big favourite to qualify... and when they lost... it was natural to seek relief under false pretences, and to escape responsibility for this failure.
"We will provide any documentation that the Asian Football Confederation wishes without wasting time."