diff --git a/apis/python/src/tiledbsoma/io/_registration/ambient_label_mappings.py b/apis/python/src/tiledbsoma/io/_registration/ambient_label_mappings.py
index 150dfaff7f..e12d971891 100644
--- a/apis/python/src/tiledbsoma/io/_registration/ambient_label_mappings.py
+++ b/apis/python/src/tiledbsoma/io/_registration/ambient_label_mappings.py
@@ -349,7 +349,10 @@ def _acquire_experiment_mappings(
     ) -> Self:
         """Acquires label-to-ID mappings from the baseline, already-written SOMA experiment."""
 
-        if experiment_uri is not None and tiledbsoma.Experiment.exists(experiment_uri):
+        if experiment_uri is not None:
+            if not tiledbsoma.Experiment.exists(experiment_uri):
+                raise ValueError("cannot find experiment at URI {experiment_uri}")
+
             # Pre-check
             with tiledbsoma.Experiment.open(experiment_uri, context=context) as exp:
                 if measurement_name not in exp.ms:
@@ -387,7 +390,10 @@ def from_anndata_appends_on_experiment(
         context: Optional[SOMATileDBContext] = None,
     ) -> Self:
         """Extends registration data from the baseline, already-written SOMA
-        experiment to include multiple H5AD input files."""
+        experiment to include multiple H5AD input files. If ``experiment_uri``
+        is ``None`` then you will be computing registrations only for the input
+        ``AnnData`` objects. If ``experiment_uri`` is not ``None`` then it is
+        an error if the experiment is not accessible."""
 
         registration_data = cls._acquire_experiment_mappings(
             experiment_uri,
diff --git a/apis/python/tests/test_registration_mappings.py b/apis/python/tests/test_registration_mappings.py
index 1419e52477..0592a2abfd 100644
--- a/apis/python/tests/test_registration_mappings.py
+++ b/apis/python/tests/test_registration_mappings.py
@@ -1164,6 +1164,23 @@ def test_ealm_expose():
     assert tiledbsoma.io.ExperimentAmbientLabelMapping is not None
 
 
+def test_append_registration_with_nonexistent_storage(tmp_path):
+    anndata1 = create_anndata_canned(1, "obs_id", "var_id")
+    anndata2 = create_anndata_canned(2, "obs_id", "var_id")
+    soma_uri = tmp_path.as_posix()
+
+    tiledbsoma.io.from_anndata(soma_uri, anndata1, measurement_name="RNA")
+
+    with pytest.raises(ValueError):
+        tiledbsoma.io.register_anndatas(
+            soma_uri + "-nonesuch",
+            [anndata2],
+            measurement_name="RNA",
+            obs_field_name="obs_id",
+            var_field_name="var_id",
+        )
+
+
 @pytest.mark.parametrize("obs_field_name", ["obs_id", "cell_id"])
 @pytest.mark.parametrize("var_field_name", ["var_id", "gene_id"])
 @pytest.mark.parametrize(