Mutual TLS won't work with HttpClientHandler

Assuming the ALB's HTTPS listener isn't set to use the TLS security policy ELBSecurityPolicy-TLS13-1-3-2021-06 that requires TLS 1.3 (https://docs.aws.amazon.com/elasticloadbalancing/latest/application/describe-ssl-policies.html#tls-protocols), it sounds likely that you may be right and that your client application just isn't presenting the client certificate when connecting to the ALB.

Are you using the same certificate, just converted with openssl between the file formats, or are these two completely separate client certificates? You also mentioned that the failing certificate/private key file is encrypted. Just to eliminate the obvious, is the client program using the passphrase to decrypt the private key?

Are you using the same certificate, just converted with openssl between the file formats, or are these two completely separate client certificates? Is the same certificate.pfx converted into 2 files *.pem 1 containing the certificate and the other containing the private key

You also mentioned that the failing certificate/private key file is encrypted. Just to eliminate the obvious, is the client program using the passphrase to decrypt the private key? With the dotnet is initialize as fallows new X509Certificat2(certificateFile, certificatePassphrase);

Tests have also been performed with curl in linux and Invoke-Webrequest in powershell and they work! So this problems is related with the dotnet libraries ....