Antibody:	alpha Enhb-1
Detection system:	Secondary antibody coupled to horseradish peroxidase
Assay notes:	Antibody used also recognizes a related gene: En2 (MGI:95390) has also been annotated.

Structure	Level	Pattern	Image	Note
TS12: neural plate	Weak	Regionally restricted	2A	Expression first observed in open neural plate as first somites were condensing. Expression rostral to pre-otic sulcus. Weak expression, relatively uniform in two patches on either side of midline. Border of patches somewhat diffuse. Expression in occasional cells not adjacent to patches. No expression in medial and lateral neuroepithelium.

Structure	Level	Pattern	Image	Note
TS12: neural plate	Strong	Regionally restricted	2B	Intense expression in every cell in region. Area of expression increased almost threefold. Rapid expansion of band due largely to rostral extension of anterior edge relative to pre-otic sulcus. No expression in medial and lateral neuroepithelium.

Structure	Level	Pattern	Image	Note
TS12: neural plate	Present	Regionally restricted	3A	Authors state expression is restricted to the neuroepithelium until the first 4 somites developed. No expression in medial and lateral neuroepithelium.

Structure	Level	Pattern	Image	Note
TS12: neural plate	Present	Regionally restricted	3B	Authors state expression is restricted to the neuroepithelium until the first 4 somites developed. No expression in medial and lateral neuroepithelium.
TS12: embryo mesenchyme	Present	Single cells	3B	Expression in a small number of cells dispersed in the mesenchyme adjacent to the edge of the neural plate.

Structure	Level	Pattern	Image	Note
TS13: migrating neural crest	Present	Regionally restricted		Expression in mesenchyme cells, possibly neural crest cells, forming a stream extending ventrally and caudally from the dorsal midline of the neural tube at the caudal edge of expressing neuroepithelium.

Structure	Level	Pattern	Image	Note
TS15: neural tube	Present	Regionally restricted	3D	Following neural tube closure, expression extends to include ventral and dorsal midlines.
TS15: midbrain-hindbrain junction	Present		3D	Expression in entire circumference of neural tube at midbrain-midbrain junction.
TS15: 1st branchial arch	Present	Regionally restricted	3D	Expression in a core of cells in the mandibular arches.

Structure	Level	Pattern	Image	Note
TS23: pituitary gland	Present	Regionally restricted	3F	Expression in cells surrounding the ventral portion of the pituitary.

Structure	Level	Pattern	Image	Note
TS12: future midbrain	Present	Regionally restricted	4A	Expression in most of the broad plate extending from just rostral to the pre-otic sulcus to the prosencephalon-mesencephalon junction.
TS12: future prosencephalon	Present	Regionally restricted	4A	Some expression also extending rostrally across the prosencephalon-mesencephalon junction.

Structure	Level	Pattern	Image	Note
TS12: future midbrain	Present	Regionally restricted	4B	Caudal limit of expression remains anterior to the pre-otic sulcus; rostral limit shifts into the mesencephalon.

Structure	Level	Pattern	Image	Note
TS13: future midbrain	Present	Regionally restricted	4C	Expression spans mesencephalic neuromere 2.
TS13: rhombomere	Present		4C	Forming rhombomeres visible caudal to the region of expression.

Structure	Level	Pattern	Image	Note
TS15: midbrain-hindbrain junction	Present			Band of expression spanning constriction at mesencephalon-metencephalon boundary.
TS15: future midbrain	Present	Regionally restricted		Rostral boundary of expression in the mesencephalon.
TS15: rhombomere 1	Present			Caudal border anterior to the rhombomere 1-rhombomere 2 border.

Structure	Level	Pattern	Image	Note
TS16: midbrain-hindbrain junction	Present		5A	Expression straddled constriction at cephalic flexure dividing mesencephalon and metencephalon.
TS16: future hindbrain	Present	Regionally restricted	5A	Expression includes a large portion of mesencephalon and most of metencephalon.
TS16: future midbrain	Present	Regionally restricted	5A	Expression includes a large portion of mesencephalon and most of metencephalon.
TS16: rhombomere 1	Present	Regionally restricted	5A	Expression ending rostral to boundary between rhombomeres 1 and 2.
TS16: future spinal cord	Present	Regionally restricted	5A	Expression extends from junction of rhombomeres 1 and 2 in the hindbrain to the caudal spinal cord.
TS16: dermatome	Present	Regionally restricted	5A	Expression in a stripe covering the lateral middle third of the dermatome portion of somites.

Structure	Level	Pattern	Image	Note
TS16: future hindbrain	Present	Regionally restricted	5B	Expression extending from junction of rhombomeres 1 and 2 in the hindbrain to the caudal spinal cord.
TS16: future spinal cord	Present	Regionally restricted	5B	Expression extending from junction of rhombomeres 1 and 2 in the hindbrain to the caudal spinal cord.
TS16: dermatome	Present	Regionally restricted	5B	Expression in a stripe covering the lateral middle third of the dermatome portion of somites. Expression at approximately the same dorsal-ventral level as expressing neurons in the spinal cord.

Structure	Level	Pattern	Image	Note
TS15: future spinal cord	Present		6A
TS15: dermatome	Present	Regionally restricted	6A	Expression in a stripe covering the lateral middle third of the dermatome portion of somites. Expression at approximately the same dorsal-ventral level as expressing neurons in the spinal cord.
TS15: forelimb bud	Present	Regionally restricted	6A	Expression first evident as a thin crescent of expressing cells along the ventral edge of the developing limb bud, preceding the formation of the apical ectodermal ridge.

Structure	Level	Pattern	Image	Note
TS14: neural tube	Present	Regionally restricted		Authors state that expression was first seen at approximately the 17 somite stage within area bounded anteriorly by otic vesicle and posteriorly by the first somite.

Structure	Level	Pattern	Image	Note
TS14: future spinal cord	Present			Expression proceeded uniformly down the length of the spinal cord.
TS14: future hindbrain	Present	Regionally restricted		Rostral progression halted just behind the otic vesicle at the junction of rhombomeres 6 and 7.

Structure	Level	Pattern	Image	Note
TS14: future spinal cord	Present
TS14: rhombomere 3	Present			Expression appeared in rhombomeres 3 and 4.
TS14: rhombomere 4	Present			Expression appeared in rhombomeres 3 and 4.

Structure	Level	Pattern	Image	Note
TS15: future spinal cord	Present
TS15: future hindbrain	Present	Regionally restricted		Final anterior limit of expression was defined at the junction between rhombomeres 1 and 2.

Structure	Level	Pattern	Image	Note
TS16: limb bud	Present	Regionally restricted		Expression in entire ventral surface of limb bud.
TS16: apical ectodermal ridge	Present	Regionally restricted		Expression in apical ridge always included the ventral portion and occasionally extended to include the dorsal ridge.
TS16: surface ectoderm	Present	Regionally restricted		Expression in ventral trunk epidermis adjacent to limb bud.

Structure	Level	Pattern	Image	Note
TS17: future spinal cord	Present		6D
TS17: dermatome	Present	Regionally restricted	6D	Expression in a stripe covering the lateral middle third of the dermatome portion of somites. Expression at approximately the same dorsal-ventral level as expressing neurons in the spinal cord.

Structure	Level	Pattern	Image	Note
TS23: spinal cord	Present	Regionally restricted	6E	Expression in cells widely spread in the ventral portion of the spinal cord.

Structure	Level	Pattern	Image	Note
TS19: forelimb bud ectoderm	Present	Regionally restricted	6F	Expression faded uniformly in the ventral ectoderm.
TS19: handplate ectoderm	Present	Regionally restricted	6F	Expression still strong in a stripe approximately 6 cells wide traversing the midline of the tips of the developing digits.