diff --git a/docs/release-notes/2399.fix.md b/docs/release-notes/2399.fix.md
new file mode 100644
index 000000000..9c3ccafc1
--- /dev/null
+++ b/docs/release-notes/2399.fix.md
@@ -0,0 +1 @@
+Disallow {meth}`anndata.AnnData.transpose` when `X` or `layers` contains {class}`h5py.Dataset`, {class}`zarr.Array` ,{class}`anndata.abc.CSRDataset`, or {class}`anndata.abc.CSCDataset` {user}`ilan-gold`.
diff --git a/src/anndata/_core/anndata.py b/src/anndata/_core/anndata.py
index 3c8574436..ae9b091be 100644
--- a/src/anndata/_core/anndata.py
+++ b/src/anndata/_core/anndata.py
@@ -1231,6 +1231,12 @@ def transpose(self) -> AnnData:
                 "which is currently not implemented. Call `.copy()` before transposing."
             )
             raise ValueError(msg)
+        if any(
+            isinstance(elem, ZarrArray | BaseCompressedSparseDataset | h5py.Dataset)
+            for elem in (self.X, *self.layers.values())
+        ):
+            msg = "Cannot transpose anndata object that has raw zarr arrays or h5py arrays backing X or layers"
+            raise ValueError(msg)
 
         return AnnData(
             X=_safe_transpose(X) if X is not None else None,
diff --git a/tests/test_base.py b/tests/test_base.py
index 2c3ec4d11..d40fcd4e7 100644
--- a/tests/test_base.py
+++ b/tests/test_base.py
@@ -5,9 +5,11 @@
 from itertools import product
 from typing import TYPE_CHECKING
 
+import h5py
 import numpy as np
 import pandas as pd
 import pytest
+import zarr
 from numpy import ma
 from scipy import sparse as sp
 from scipy.sparse import csr_matrix, issparse
@@ -15,6 +17,7 @@
 import anndata as ad
 from anndata import AnnData, ImplicitModificationWarning
 from anndata._core.raw import Raw
+from anndata._core.sparse_dataset import sparse_dataset
 from anndata._settings import settings
 from anndata.acc import A
 from anndata.tests.helpers import (
@@ -834,3 +837,23 @@ def test_create_adata_from_single_axis_elem(
     in_memory.write_h5ad(tmp_path / "adata.h5ad")
     from_disk = ad.read_h5ad(tmp_path / "adata.h5ad")
     assert_equal(from_disk, in_memory)
+
+
+@pytest.mark.parametrize("in_x", [True, False], ids=["X", "layers"])
+@pytest.mark.parametrize("is_sparse", [True, False], ids=["sparse", "dense"])
+@pytest.mark.parametrize("storage", ["h5ad", "zarr"])
+def test_transpose_errors_with_backed_arrays(
+    tmp_path: Path, storage: str, *, is_sparse: bool, in_x: bool
+):
+    adata = AnnData(X=csr_matrix(np.ones((3, 4))) if is_sparse else np.ones((3, 4)))
+    path = tmp_path / f"test.{storage}"
+    getattr(adata, f"write_{storage}")(path)
+    f = (h5py.File if storage == "h5ad" else zarr.open)(path)
+    raw_array = sparse_dataset(f["X"]) if is_sparse else f["X"]
+
+    adata = AnnData(**({"X": raw_array} if in_x else {"layers": {"test": raw_array}}))
+
+    with pytest.raises(ValueError, match=r"Cannot transpose anndata object"):
+        adata.transpose()
+    if storage == "h5ad":
+        f.close()