China ordered all domestic organizations and individuals to cease cooperation with a European Union antitrust investigation into JD.com.
The Ministry of Justice described the EU probe as improper extraterritorial jurisdiction and a violation of international law.
The EU launched the inquiry in May under the Foreign Subsidies Regulation.
Regulators are investigating whether state subsidies provided JD.com an unfair advantage in its $2.5 billion acquisition of German retailer Ceconomy.
This marks the second time Beijing has used counter-sanction laws to block an EU corporate investigation.
The move signals an intensifying regulatory and trade clash between the two powers.