The Reds were expected to have a near-fully fit first-team squad to face the Saints with Sadio Mané also expected to be made available to feature against his former club.
The only absentee would have been right-back Nathaniel Clyne, with Adam Lallana, Jordan Henderson and Philippe Coutinho all fit to be involved and Mané likely to make the match-day squad.
But Matip's injury - the news of which only broke late on Friday night - puts a dent into Klopp's plans with Dejan Lovren either set to be recalled or Joe Gomez to be repositioned.
Matip has reportedly been struggling with an adductor muscle issue and is not expected to be risked against Mauricio Pochettino's visitors.
Gomez has started all of his outings this season at right-back, rotating with Trent Alexander-Arnold with Clyne sidelined, but impressed centrally in his first couple of caps for England this week.
Lovren, meanwhile, has not started since being withdrawn after a disastrous opening half-an-hour in Liverpool's 4-1 defeat at Tottenham Hotspur.
The Croatian desperately struggled up against Harry Kane and was responsible in part for both of Spurs' opening two goals, that gave them a strong advantage after just 12 minutes at Wembley Stadium.
Lovren did complete 90 minutes twice for Croatia over the international break as his country beat Greece 4-1 over two legs in a World Cup play-off to book their place in next summer's finals.
That could tempt to Klopp to restore the 28-year-old alongside Ragnar Klavan at centre-back, with Alberto Moreno a certainty to start at left-back and either Gomez or Alexander-Arnold starting on the right.
Yet it is an unnecessary headache that Klopp could do without as he prepares to face Southampton, looking to mastermind his side to their fourth straight win in all competitions since the Spurs debacle.
Matip's untimely absence a blow to Reds
Matip has developed a worrying tendency to pick up niggling injuries, missing 10 games last term due to ankle problems, and Klopp will hope this issue is not a precursor for more minor issues to come ahead of a busy fixture schedule.
It is even more of an issue given that Matip is the club's most dependable senior centre-back, having been picked whenever fit since signing from Schalke in the summer of 2016.
The Cameroonian also netted in Liverpool's last outing before the international break, firing in a rebound at a corner for his first goal in a year - although he has proven a threat, even if an unprolific one, at set-pieces this term.
Gomez's promise is obvious but he has played few games in the middle since joining from Charlton Athletic in the summer of 2015 and moving him inside leaves the problem of having just one right-back in youngster Alexander-Arnold.
It is therefore heavily likely that Lovren will be recalled and with it inevitable he will be met with an irate reception from Southampton fans after the way he left the club in order to sign for Liverpool in 2014, his reception from the home fans may also be mixed.
Lovren's inconsistency and propensity to commit basic errors has seen him repeatedly come under fire during his Anfield career, with the player voted as the club's Player of the Month for October in a bid to boost his morale following a tirade of abuse on social media.
Liverpool face 13 Premier League and Champions League matches across the next 45 days, an average of a game every three days, and will surely have to lend on Matip heavily if they are to maintain defensive solidity and put together a consistent run of results.
Given how soft Liverpool's centre has been shown to be by several teams, and how important a fit and in-form Matip is to their back-line, the Merseyside outfit will hope his adductor issue is only a minor one.