diff --git a/packages/vite/src/node/preview.ts b/packages/vite/src/node/preview.ts
index d17fc0845326dc..8148b34110022b 100644
--- a/packages/vite/src/node/preview.ts
+++ b/packages/vite/src/node/preview.ts
@@ -15,7 +15,7 @@ import {
 import { openBrowser } from './server/openBrowser'
 import compression from './server/middlewares/compression'
 import { proxyMiddleware } from './server/middlewares/proxy'
-import { resolveHostname, resolveServerUrls } from './utils'
+import { resolveHostname, resolveServerUrls, shouldServe } from './utils'
 import { printServerUrls } from './logger'
 import { resolveConfig } from '.'
 import type { InlineConfig, ResolvedConfig } from '.'
@@ -112,21 +112,24 @@ export async function preview(
   // static assets
   const distDir = path.resolve(config.root, config.build.outDir)
   const headers = config.preview.headers
-  app.use(
-    previewBase,
-    sirv(distDir, {
-      etag: true,
-      dev: true,
-      single: config.appType === 'spa',
-      setHeaders(res) {
-        if (headers) {
-          for (const name in headers) {
-            res.setHeader(name, headers[name]!)
-          }
+  const assetServer = sirv(distDir, {
+    etag: true,
+    dev: true,
+    single: config.appType === 'spa',
+    setHeaders(res) {
+      if (headers) {
+        for (const name in headers) {
+          res.setHeader(name, headers[name]!)
         }
       }
-    })
-  )
+    }
+  })
+  app.use(previewBase, async (req, res, next) => {
+    if (shouldServe(req.url!, distDir)) {
+      return assetServer(req, res, next)
+    }
+    next()
+  })
 
   // apply post server hooks from plugins
   postHooks.forEach((fn) => fn && fn())
diff --git a/packages/vite/src/node/server/middlewares/static.ts b/packages/vite/src/node/server/middlewares/static.ts
index c877022a63cde7..8f6b47f3a7e4bb 100644
--- a/packages/vite/src/node/server/middlewares/static.ts
+++ b/packages/vite/src/node/server/middlewares/static.ts
@@ -14,6 +14,7 @@ import {
   isInternalRequest,
   isParentDirectory,
   isWindows,
+  shouldServe,
   slash
 } from '../../utils'
 
@@ -52,7 +53,10 @@ export function servePublicMiddleware(
     if (isImportRequest(req.url!) || isInternalRequest(req.url!)) {
       return next()
     }
-    serve(req, res, next)
+    if (shouldServe(req.url!, dir)) {
+      return serve(req, res, next)
+    }
+    next()
   }
 }
 
diff --git a/packages/vite/src/node/utils.ts b/packages/vite/src/node/utils.ts
index 23c7597bb77195..3c1e9c90f2a034 100644
--- a/packages/vite/src/node/utils.ts
+++ b/packages/vite/src/node/utils.ts
@@ -1198,6 +1198,45 @@ export const isNonDriveRelativeAbsolutePath = (p: string): boolean => {
   return windowsDrivePathPrefixRE.test(p)
 }
 
+/**
+ * Determine if a file is being requested with the correct case, to ensure
+ * consistent behaviour between dev and prod and across operating systems.
+ */
+export function shouldServe(url: string, assetsDir: string): boolean {
+  // viteTestUrl is set to something like http://localhost:4173/ and then many tests make calls
+  // like `await page.goto(viteTestUrl + '/example')` giving us URLs beginning with a double slash
+  const pathname = decodeURI(
+    new URL(url.startsWith('//') ? url.substring(1) : url, 'http://example.com')
+      .pathname
+  )
+  const file = path.join(assetsDir, pathname)
+  if (
+    !fs.existsSync(file) ||
+    (isCaseInsensitiveFS && // can skip case check on Linux
+      !fs.statSync(file).isDirectory() &&
+      !hasCorrectCase(file, assetsDir))
+  ) {
+    return false
+  }
+  return true
+}
+
+/**
+ * Note that we can't use realpath here, because we don't want to follow
+ * symlinks.
+ */
+function hasCorrectCase(file: string, assets: string): boolean {
+  if (file === assets) return true
+
+  const parent = path.dirname(file)
+
+  if (fs.readdirSync(parent).includes(path.basename(file))) {
+    return hasCorrectCase(parent, assets)
+  }
+
+  return false
+}
+
 export function joinUrlSegments(a: string, b: string): string {
   if (!a || !b) {
     return a || b || ''
diff --git a/playground/assets/__tests__/assets.spec.ts b/playground/assets/__tests__/assets.spec.ts
index 768fadf7f35ce5..f8246a4f3418b3 100644
--- a/playground/assets/__tests__/assets.spec.ts
+++ b/playground/assets/__tests__/assets.spec.ts
@@ -1,3 +1,4 @@
+import fetch from 'node-fetch'
 import { describe, expect, test } from 'vitest'
 import {
   browserLogs,
@@ -12,6 +13,7 @@ import {
   readFile,
   readManifest,
   untilUpdated,
+  viteTestUrl,
   watcher
 } from '~utils'
 
@@ -27,6 +29,11 @@ test('should have no 404s', () => {
   })
 })
 
+test('should get a 404 when using incorrect case', async () => {
+  expect((await fetch(viteTestUrl + 'icon.png')).status).toBe(200)
+  expect((await fetch(viteTestUrl + 'ICON.png')).status).toBe(404)
+})
+
 describe('injected scripts', () => {
   test('@vite/client', async () => {
     const hasClient = await page.$(