Amyloid can be suspected on H&E stain as a lightly eosinophilic, amorphous, “smudgy” deposit seen particularly within arterioles. The deposits often obscure the nuclei of arteriole smooth muscle cells. Occasionally, the degree of amyloid deposition is more advanced with involvement beyond the arteriole and infiltration into the bone marrow interstitium. In a few cases, the extent of amyloid deposition is so severe with geographic deposits replacing entire intertrabecular spaces. Interstitial amyloid deposits, especially when they are not associated with a vessel, should be differentiated from “bone dust” (an artifact seen as the result of the bone marrow biopsy procedure), fibrin, and serous fat atrophy.

Congo red is the most specific stain for amyloid and yields the characteristic orange/pink or “salmon” color [2]. We have devised a scale to convey the presence and degree of amyloid deposition that ranges from no amyloid deposition (0), amyloid localized to blood vessels (1+), and involvement of the interstitium less than two high power fields in size (2+) or greater than two high power fields in size (3+) (Fig. 26.2) [3]. For gold standard confirmation, the Congo red stain demonstrates the characteristic “apple green” birefringence when viewed under polarized light (Fig. 26.3) [2]. Congo red can be particularly helpful in detecting amyloid deposition in the periosteum, where clearance likely takes longer compared to the bone marrow and other organs. Mimics such as “bone dust” can appear Congophilic; however, it will not demonstrate “apple green” birefringence with polarized light. Also of note, not all birefringence is the same, for example collagen fibers in lamellar bone will emit a “silvery” birefringence in contrast to the “apple green” birefringence seen in amyloid.

In our case series of 462 bone marrow biopsies from AL amyloidosis patients, 268 had amyloid deposition in the bone marrow whereas 194 did not (Table 26.1). Of the 268 bone marrows showing demonstrable amyloid involvement, 74 % had 1+, 15 % had 2+, and 11 % had 3+ involvement. We have come to find that this grading scale has no predictive ability for hematologic status, bone marrow stem cell mobilization after administering colony stimulating factor, bone marrow engraftment after ablative chemotherapy, or degree of involvement or function in other organs. Lastly, bone marrow amyloid deposition can be seen in non-AL amyloidosis such as AA type amyloidosis [3]. Hence, finding amyloid deposition in the bone marrow is usually, but not always, attributable to an amyloidogenic plasma cell clone (AL amyloidosis).

In AL amyloidosis, demonstrating the clonality [1, 4–6] and measuring the volume [5, 6] of plasma cells are the most important parameters from the bone marrow biopsy in facilitating the diagnosis [1, 4–6] and predicting prognosis [5, 6], respectively. Plasma cells can be identified on H&E stain, but precise enumeration is difficult especially when dealing with small volumes. Unlike multiple myeloma, the plasma cell volume in AL amyloidosis is often low and is on average around 10 % of all nucleated cells (Table 26.1). Our ability to enumerate plasma cells in the face of such low volumes has been greatly enhanced with the use of IHC directed at the CD138 antigen, a marker specific for plasma cells in the bone marrow (Fig. 26.4). IHC with CD138 highlights all plasma cells including the amyloidogenic clone and normal plasma cells. Rarely, amyloid may show nonspecific “sticky” staining with CD138 IHC, which may make plasma cell enumeration difficult especially when amyloid deposits are geographic. As an alternative, MUM1 IHC can be used in these situations in enumerating plasma cells. Of the few cases in which we have used MUM1, the amyloid deposits have not shown nonspecific staining.

Identification of a plasma cell clone requires the demonstration of either kappa or lambda light chain predominance. Plasma cells can only express a single light chain and there are up to three times as many kappa expressing plasma cells as lambda (which is applicable to both bone marrow and extramedullary sites). The normal kappa:lambda ratio is 1–3:1 based on the fact that a B-cell will try to rearrange both kappa light chain genes before attempting to rearrange either of the lambda light chain genes. A mixture of plasma cells with a kappa:lambda ratio of 1–3:1 is presumably polyclonal. Light chain predominance is seen when either the kappa:lambda ratio exceeds 3:1 or a kappa:lambda ratio is less than 1:1, denoting either kappa or lambda light chain predominance, respectively [1]. Once light chain predominance is established, it can then be presumed that there is a light chain-restricted clone of the same type upsetting the normal kappa:lambda ratio.

Kappa and lambda light chains can either be stained by targeting the protein (IHC) or the messenger RNA (colorimetric in situ hybridization or CISH). The favored modality of light chain staining is CISH rather than IHC (Fig. 26.5) [7]. In CISH, oligonucleotide probes are directed against the conserved regions of the kappa and lambda light chain messenger RNA [7]. These probes are coupled to a chromogen, which is visualized as blue-black pigment [7]. IHC against the kappa and lambda light chain protein is fraught with nonspecific staining, especially for formalin-based fixatives [1, 7]. Determination of either kappa or lambda light chain predominance requires a tandem comparison of the cellular staining volume seen with kappa versus lambda. In rare cases, amyloid may show dual nonspecific “sticky” staining for both kappa and lambda CISH. If there are geographic amyloid deposits showing nonspecific staining for both kappa and lambda CISH, demonstrating light chain predominance can be difficult or even impossible.

Of the 462 bone marrow biopsies from newly diagnosed and untreated patients with AL amyloidosis who have had clonality studies done in our clinic, 88 % showed a demonstrable plasma cell neoplasm compared to 12 % in which a plasma cell neoplasm was not demonstrable by bone marrow biopsy (Table 26.1). Of the 88 % with a demonstrable plasma cell neoplasm, 66 % were lambda-restricted and 22 % were kappa-restricted, a finding that was consistent with other published reports [4]. An example of a pre- and posttreatment bone marrow evaluation from a patient is shown in Fig. 26.6. The pretreatment bone marrow clearly shows a lambda-restricted plasma cell clone, whereas the posttreatment bone marrow shows polyclonal staining.